Process definitions¶

ATAC-Seq¶

data:workflow:atacseqworkflow-atac-seq (data:reads:fastq reads, data:index:bowtie2 genome, data:bed promoter, basic:string mode, basic:string speed, basic:boolean use_se, basic:boolean discordantly, basic:boolean rep_se, basic:integer minins, basic:integer maxins, basic:integer trim_5, basic:integer trim_3, basic:integer trim_iter, basic:integer trim_nucl, basic:string rep_mode, basic:integer k_reports, basic:integer q_threshold, basic:integer n_sub, basic:boolean tn5, basic:integer shift, basic:boolean tagalign, basic:string duplicates, basic:string duplicates_prepeak, basic:decimal qvalue, basic:decimal pvalue, basic:decimal pvalue_prepeak, basic:integer cap_num, basic:integer mfold_lower, basic:integer mfold_upper, basic:integer slocal, basic:integer llocal, basic:integer extsize, basic:integer shift, basic:integer band_width, basic:boolean nolambda, basic:boolean fix_bimodal, basic:boolean nomodel, basic:boolean nomodel_prepeak, basic:boolean down_sample, basic:boolean bedgraph, basic:boolean spmr, basic:boolean call_summits, basic:boolean broad, basic:decimal broad_cutoff)[Source: v3.0.1]

This ATAC-seq pipeline closely follows the official ENCODE DCC pipeline. It is comprised of three steps; alignment, pre-peakcall QC, and calling peaks (with post-peakcall QC). First, reads are aligned to a genome using [Bowtie2](http://bowtie-bio.sourceforge.net/index.shtml) aligner. Next, pre-peakcall QC metrics are calculated. QC report contains ENCODE 3 proposed QC metrics – [NRF](https://www.encodeproject.org/data-standards/terms/), [PBC bottlenecking coefficients, NSC, and RSC](https://genome.ucsc.edu/ENCODE/qualityMetrics.html#chipSeq). Finally, the peaks are called using [MACS2](https://github.com/taoliu/MACS/). The post-peakcall QC report includes additional QC metrics – number of peaks, fraction of reads in peaks (FRiP), number of reads in peaks, and if promoter regions BED file is provided, number of reads in promoter regions, fraction of reads in promoter regions, number of peaks in promoter regions, and fraction of reads in promoter regions.

reads

label: Select sample(s)
type: data:reads:fastq

genome

label: Genome
type: data:index:bowtie2

promoter

label: Promoter regions BED file
type: data:bed
description: BED file containing promoter regions (TSS+-1000 bp for example). Needed to get the number of peaks and reads mapped to promoter regions.
required: False

alignment.mode

label

Alignment mode

type

basic:string

description

End to end: Bowtie 2 requires that the entire read align from one end to the other, without any trimming (or “soft clipping”) of characters from either end. Local: Bowtie 2 does not require that the entire read align from one end to the other. Rather, some characters may be omitted (“soft clipped”) from the ends in order to achieve the greatest possible alignment score.

default

--local

choices

end to end mode: --end-to-end
local: --local

alignment.speed

label

Speed vs. Sensitivity

type

basic:string

default

--sensitive

choices

Very fast: --very-fast
Fast: --fast
Sensitive: --sensitive
Very sensitive: --very-sensitive

alignment.PE_options.use_se

label: Map as single-ended (for paired-end reads only)
type: basic:boolean
description: If this option is selected paired-end reads will be mapped as single-ended and other paired-end options are ignored.
default: False

alignment.PE_options.discordantly

label: Report discordantly matched read
type: basic:boolean
description: If both mates have unique alignments, but the alignments do not match paired-end expectations (orientation and relative distance) then alignment will be reported. Useful for detecting structural variations.
default: True

alignment.PE_options.rep_se

label: Report single ended
type: basic:boolean
description: If paired alignment can not be found Bowtie2 tries to find alignments for the individual mates.
default: True

alignment.PE_options.minins

label: Minimal distance
type: basic:integer
description: The minimum fragment length for valid paired-end alignments. 0 imposes no minimum.
default: 0

alignment.PE_options.maxins

label: Maximal distance
type: basic:integer
description: The maximum fragment length for valid paired-end alignments.
default: 2000

alignment.start_trimming.trim_5

label: Bases to trim from 5’
type: basic:integer
description: Number of bases to trim from from 5’ (left) end of each read before alignment.
default: 0

alignment.start_trimming.trim_3

label: Bases to trim from 3’
type: basic:integer
description: Number of bases to trim from from 3’ (right) end of each read before alignment
default: 0

alignment.trimming.trim_iter

label: Iterations
type: basic:integer
description: Number of iterations.
default: 0

alignment.trimming.trim_nucl

label: Bases to trim
type: basic:integer
description: Number of bases to trim from 3’ end in each iteration.
default: 2

alignment.reporting.rep_mode

label

Report mode

type

basic:string

description

Default mode: search for multiple alignments, report the best one; -k mode: search for one or more alignments, report each; -a mode: search for and report all alignments

default

def

choices

Default mode: def
-k mode: k
-a mode (very slow): a

alignment.reporting.k_reports

label: Number of reports (for -k mode only)
type: basic:integer
description: Searches for at most X distinct, valid alignments for each read. The search terminates when it can’t find more distinct valid alignments, or when it finds X, whichever happens first.
default: 5

prepeakqc_settings.q_threshold

label: Quality filtering threshold
type: basic:integer
default: 30

prepeakqc_settings.n_sub

label: Number of reads to subsample
type: basic:integer
default: 25000000

prepeakqc_settings.tn5

label: Tn5 shifting
type: basic:boolean
description: Tn5 transposon shifting. Shift reads on “+” strand by 4 bp and reads on “-” strand by 5 bp.
default: True

prepeakqc_settings.shift

label: User-defined cross-correlation peak strandshift
type: basic:integer
description: If defined, SPP tool will not try to estimate fragment length but will use the given value as fragment length.
default: 0

settings.tagalign

label: Use tagAlign files
type: basic:boolean
description: Use filtered tagAlign files as case (treatment) and control (background) samples. If extsize parameter is not set, run MACS using input’s estimated fragment length.
default: True

settings.duplicates

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

settings.tagalign

choices

1: 1
auto: auto
all: all

settings.duplicates_prepeak

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

!settings.tagalign

default

all

choices

1: 1
auto: auto
all: all

settings.qvalue

label: Q-value cutoff
type: basic:decimal
description: The q-value (minimum FDR) cutoff to call significant regions. Q-values are calculated from p-values using Benjamini-Hochberg procedure.
required: False
disabled: settings.pvalue && settings.pvalue_prepeak

settings.pvalue

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
required: False
disabled: settings.qvalue
hidden: settings.tagalign

settings.pvalue_prepeak

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
disabled: settings.qvalue
hidden: !settings.tagalign || settings.qvalue
default: 0.01

settings.cap_num

label: Cap number of peaks by taking top N peaks
type: basic:integer
description: To keep all peaks set value to 0.
disabled: settings.broad
default: 300000

settings.mfold_lower

label: MFOLD range (lower limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.mfold_upper

label: MFOLD range (upper limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.slocal

label: Small local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.llocal

label: Large local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.extsize

label: extsize
type: basic:integer
description: While ‘–nomodel’ is set, MACS uses this parameter to extend reads in 5’->3’ direction to fix-sized fragments. For example, if the size of binding region for your transcription factor is 200 bp, and you want to bypass the model building by MACS, this parameter can be set as 200. This option is only valid when –nomodel is set or when MACS fails to build model and –fix-bimodal is on.
default: 150

settings.shift

label: Shift
type: basic:integer
description: Note, this is NOT the legacy –shiftsize option which is replaced by –extsize! You can set an arbitrary shift in bp here. Please Use discretion while setting it other than default value (0). When –nomodel is set, MACS will use this value to move cutting ends (5’) then apply –extsize from 5’ to 3’ direction to extend them to fragments. When this value is negative, ends will be moved toward 3’->5’ direction, otherwise 5’->3’ direction. Recommended to keep it as default 0 for ChIP-Seq datasets, or -1 * half of EXTSIZE together with –extsize option for detecting enriched cutting loci such as certain DNAseI-Seq datasets. Note, you can’t set values other than 0 if format is BAMPE for paired-end data. Default is 0.
default: -75

settings.band_width

label: Band width
type: basic:integer
description: The band width which is used to scan the genome ONLY for model building. You can set this parameter as the sonication fragment size expected from wet experiment. The previous side effect on the peak detection process has been removed. So this parameter only affects the model building.
required: False

settings.nolambda

label: Use backgroud lambda as local lambda
type: basic:boolean
description: With this flag on, MACS will use the background lambda as local lambda. This means MACS will not consider the local bias at peak candidate regions.
default: False

settings.fix_bimodal

label: Turn on the auto paired-peak model process
type: basic:boolean
description: Turn on the auto paired-peak model process. If it’s set, when MACS failed to build paired model, it will use the nomodel settings, the ‘–extsize’ parameter to extend each tag. If set, MACS will be terminated if paired-peak model has failed.
default: False

settings.nomodel

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: settings.tagalign
default: False

settings.nomodel_prepeak

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: !settings.tagalign
default: True

settings.down_sample

label: Down-sample
type: basic:boolean
description: When set to true, random sampling method will scale down the bigger sample. By default, MACS uses linear scaling. This option will make the results unstable and irreproducible since each time, random reads would be selected, especially the numbers (pileup, pvalue, qvalue) would change.
default: False

settings.bedgraph

label: Save fragment pileup and control lambda
type: basic:boolean
description: If this flag is on, MACS will store the fragment pileup, control lambda, -log10pvalue and -log10qvalue scores in bedGraph files. The bedGraph files will be stored in current directory named NAME+’_treat_pileup.bdg’ for treatment data, NAME+’_control_lambda.bdg’ for local lambda values from control, NAME+’_treat_pvalue.bdg’ for Poisson pvalue scores (in -log10(pvalue) form), and NAME+’_treat_qvalue.bdg’ for q-value scores from Benjamini-Hochberg-Yekutieli procedure.
default: True

settings.spmr

label: Save signal per million reads for fragment pileup profiles
type: basic:boolean
disabled: settings.bedgraph === false
default: True

settings.call_summits

label: Call summits
type: basic:boolean
description: MACS will now reanalyze the shape of signal profile (p or q-score depending on cutoff setting) to deconvolve subpeaks within each peak called from general procedure. It’s highly recommended to detect adjacent binding events. While used, the output subpeaks of a big peak region will have the same peak boundaries, and different scores and peak summit positions.
default: True

settings.broad

label: Composite broad regions
type: basic:boolean
description: When this flag is on, MACS will try to composite broad regions in BED12 (a gene-model-like format) by putting nearby highly enriched regions into a broad region with loose cutoff. The broad region is controlled by another cutoff through –broad-cutoff. The maximum length of broad region length is 4 times of d from MACS.
disabled: settings.call_summits === true
default: False

settings.broad_cutoff

label: Broad cutoff
type: basic:decimal
description: Cutoff for broad region. This option is not available unless –broad is set. If -p is set, this is a p-value cutoff, otherwise, it’s a q-value cutoff. DEFAULT = 0.1
required: False
disabled: settings.call_summits === true || settings.broad !== true

Abstract alignment process¶

data:alignmentabstract-alignment ()[Source: v1.0.0]

bam

label: Alignment file
type: basic:file

bai

label: Alignment index BAI
type: basic:file

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Abstract annotation process¶

data:annotationabstract-annotation ()[Source: v1.0.0]

annot

label: Uploaded file
type: basic:file

source

label: Gene ID source
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Abstract bed process¶

data:bedabstract-bed ()[Source: v1.0.1]

bed

label: BED
type: basic:file

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Abstract differential expression process¶

data:differentialexpressionabstract-differentialexpression ()[Source: v1.0.0]

raw

label: Differential expression (gene level)
type: basic:file

de_json

label: Results table (JSON)
type: basic:json

de_file

label: Results table (file)
type: basic:file

source

label: Gene ID source
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

feature_type

label: Feature type
type: basic:string

Abstract expression process¶

data:expressionabstract-expression ()[Source: v1.0.0]

exp

label: Normalized expression
type: basic:file

rc

label: Read counts
type: basic:file
required: False

exp_json

label: Expression (json)
type: basic:json

exp_type

label: Expression type
type: basic:string

source

label: Gene ID source
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

feature_type

label: Feature type
type: basic:string

Accel Amplicon Pipeline¶

data:workflow:ampliconworkflow-accel (data:reads:fastq:paired reads, data:seq:nucleotide genome, data:index:bwa bwa_index, data:masterfile:amplicon master_file, data:seq:nucleotide adapters, list:data:variants:vcf known_indels, list:data:variants:vcf known_vars, data:variants:vcf dbsnp, basic:integer mbq, basic:integer stand_call_conf, basic:integer min_bq, basic:integer min_alt_bq, list:data:variants:vcf known_vars_db, basic:decimal af_threshold)[Source: v5.0.1]

Processing pipeline to analyse the Accel-Amplicon NGS panel data. The raw amplicon sequencing reads are quality trimmed using Trimmomatic. The quality of the raw and trimmed data is assesed using the FASTQC tool. Quality trimmed reads are aligned to a reference genome using BWA mem. Sequencing primers are removed from the aligned reads using Primerclip. Amplicon performance stats are calculated using Bedtools coveragebed and Picard CollectTargetedPcrMetrics programs. Prior to variant calling, the alignment file is preprocessed using the GATK IndelRealigner and BaseRecalibrator tools. GATK HaplotypeCaller and Lofreq tools are used to call germline variants. Called variants are annotated using the SnpEff tool. Finally, the amplicon performance metrics and identified variants data are used to generate the PDF analysis report.

reads

label: Input reads
type: data:reads:fastq:paired

genome

label: Genome sequence (FASTA)
type: data:seq:nucleotide

bwa_index

label: Genome index (BWA)
type: data:index:bwa

master_file

label: Experiment Master file
type: data:masterfile:amplicon

adapters

label: Adapters
type: data:seq:nucleotide
description: Provide an Illumina sequencing adapters file (.fasta) with adapters to be removed by Trimmomatic.

preprocess_bam.known_indels

label: Known indels
type: list:data:variants:vcf

preprocess_bam.known_vars

label: Known variants
type: list:data:variants:vcf

gatk.dbsnp

label: dbSNP
type: data:variants:vcf

gatk.mbq

label: Min Base Quality
type: basic:integer
description: Minimum base quality required to consider a base for calling.
default: 20

gatk.stand_call_conf

label: Min call confidence threshold
type: basic:integer
description: The minimum phred-scaled confidence threshold at which variants should be called.
default: 20

lofreq.min_bq

label: Min baseQ
type: basic:integer
description: Skip any base with baseQ smaller than the default value.
default: 20

lofreq.min_alt_bq

label: Min alternate baseQ
type: basic:integer
description: Skip alternate bases with baseQ smaller than the default value.
default: 20

var_annot.known_vars_db

label: Known variants
type: list:data:variants:vcf

report.af_threshold

label: Allele frequency threshold
type: basic:decimal
default: 0.01

Align (BWA) and trim adapters¶

data:alignment:bam:bwatrimalign-bwa-trim (data:masterfile:amplicon master_file, data:index:bwa genome, data:reads:fastq reads, basic:integer seed_l, basic:integer band_w, basic:decimal re_seeding, basic:boolean m, basic:integer match, basic:integer missmatch, basic:integer gap_o, basic:integer gap_e, basic:integer clipping, basic:integer unpaired_p, basic:boolean report_all, basic:integer report_tr)[Source: v2.1.1]

Align with BWA mem and trim the sam output. The process uses the memory-optimized Primertrim tool.

master_file

label: Master file
type: data:masterfile:amplicon
description: Amplicon experiment design file that holds the information about the primers to be removed.

genome

label: Reference genome
type: data:index:bwa

reads

label: Reads
type: data:reads:fastq

seed_l

label: Minimum seed length
type: basic:integer
description: Minimum seed length. Matches shorter than minimum seed length will be missed. The alignment speed is usually insensitive to this value unless it significantly deviates 20.
default: 19

band_w

label: Band width
type: basic:integer
description: Gaps longer than this will not be found.
default: 100

re_seeding

label: Re-seeding factor
type: basic:decimal
description: Trigger re-seeding for a MEM longer than minSeedLen*FACTOR. This is a key heuristic parameter for tuning the performance. Larger value yields fewer seeds, which leads to faster alignment speed but lower accuracy.
default: 1.5

m

label: Mark shorter split hits as secondary
type: basic:boolean
description: Mark shorter split hits as secondary (for Picard compatibility)
default: False

scoring.match

label: Score of a match
type: basic:integer
default: 1

scoring.missmatch

label: Mismatch penalty
type: basic:integer
default: 4

scoring.gap_o

label: Gap open penalty
type: basic:integer
default: 6

scoring.gap_e

label: Gap extension penalty
type: basic:integer
default: 1

scoring.clipping

label: Clipping penalty
type: basic:integer
description: Clipping is applied if final alignment score is smaller than (best score reaching the end of query) - (Clipping penalty)
default: 5

scoring.unpaired_p

label: Penalty for an unpaired read pair
type: basic:integer
description: Affinity to force pair. Score: scoreRead1+scoreRead2-Penalty
default: 9

reporting.report_all

label: Report all found alignments
type: basic:boolean
description: Output all found alignments for single-end or unpaired paired-end reads. These alignments will be flagged as secondary alignments.
default: False

reporting.report_tr

label: Report threshold score
type: basic:integer
description: Don’t output alignment with score lower than defined number. This option only affects output.
default: 30

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Alleyoop UTR Rates¶

data:alleyoop:utrrates:alleyoop-utr-rates (data:seq:nucleotide ref_seq, data:bed regions, data:alignment:bam:slamdunk slamdunk, basic:integer read_length)[Source: v1.2.1]

Run Alleyoop utrrates.

ref_seq

label: FASTA file containig sequences for aligning
type: data:seq:nucleotide
required: True
hidden: False

regions

label: BED file with coordinates of regions of interest
type: data:bed
required: True
hidden: False

slamdunk

label: Slamdunk results
type: data:alignment:bam:slamdunk
required: True
hidden: False

read_length

label: Maximum read length
type: basic:integer
description: Maximum length of reads in the input FASTQ file
required: True
hidden: False
default: 150

report

label: Tab-separated file containing conversion rates on each region of interest
type: basic:file
required: True
hidden: False

plot

label: Region of interest conversion rate plot
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Alleyoop collapse¶

data:alleyoop:collapse:alleyoop-collapse (data:alignment:bam:slamdunk slamdunk, basic:string source)[Source: v1.2.1]

Run Alleyoop collapse tool on Slamdunk results.

slamdunk

label: Slamdunk results
type: data:alignment:bam:slamdunk
required: True
hidden: False

source

label

Gene ID source

type

basic:string

required

True

hidden

False

default

ENSEMBL

choices

ENSEMBL: ENSEMBL
UCSC: UCSC

tcount

label: Count report containing SLAMSeq statistics
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Alleyoop rates¶

data:alleyoop:rates:alleyoop-rates (data:seq:nucleotide ref_seq, data:alignment:bam:slamdunk slamdunk)[Source: v1.1.1]

Run Alleyoop rates.

ref_seq

label: FASTA file containig sequences for aligning
type: data:seq:nucleotide
required: True
hidden: False

slamdunk

label: Slamdunk results
type: data:alignment:bam:slamdunk
required: True
hidden: False

report

label: Tab-separated file containing the overall conversion rates
type: basic:file
required: True
hidden: False

plot

label: Overall conversion rate plot file
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Alleyoop snpeval¶

data:alleyoop:snpeval:alleyoop-snpeval (data:seq:nucleotide ref_seq, data:bed regions, data:alignment:bam:slamdunk slamdunk, basic:integer read_length)[Source: v1.2.1]

Run Alleyoop snpeval.

ref_seq

label: FASTA file containig sequences for aligning
type: data:seq:nucleotide
required: True
hidden: False

regions

label: BED file with coordinates of regions of interest
type: data:bed
required: True
hidden: False

slamdunk

label: Slamdunk results
type: data:alignment:bam:slamdunk
required: True
hidden: False

read_length

label: Maximum read length
type: basic:integer
description: Maximum length of reads in the input FASTQ file
required: True
hidden: False
default: 150

report

label: Tab-separated file with read counts, T>C read counts and SNP indication
type: basic:file
required: True
hidden: False

plot

label: SNP evaluation plot
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Alleyoop summary¶

data:alleyoop:summary:alleyoop-summary (list:data:alignment:bam:slamdunk slamdunk)[Source: v1.1.1]

Run Alleyoop summary.

slamdunk

label: Slamdunk results
type: list:data:alignment:bam:slamdunk
required: True
hidden: False

report

label: Tab-separated file with mapping statistics
type: basic:file
required: True
hidden: False

plot_data

label: PCA values of the samples based on T>C read counts in regions of interest.
type: basic:file
required: False
hidden: False

plot

label: PCA plot of the samples based on T>C read counts in regions of interest.
type: basic:file
required: False
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Amplicon report¶

data:report:ampliconamplicon-report (data:picard:coverage pcr_metrics, data:coverage coverage, data:masterfile:amplicon master_file, list:data:snpeff annot_vars, basic:decimal af_threshold)[Source: v1.1.1]

Create amplicon report.

pcr_metrics

label: Picard TargetedPcrMetrics
type: data:picard:coverage

coverage

label: Coverage
type: data:coverage

master_file

label: Amplicon master file
type: data:masterfile:amplicon

annot_vars

label: Annotated variants (snpEff)
type: list:data:snpeff

af_threshold

label: Allele frequency threshold
type: basic:decimal
default: 0.01

report

label: Report
type: basic:file

panel_name

label: Panel name
type: basic:string

stats

label: File with sample statistics
type: basic:file

amplicon_cov

label: Amplicon coverage file (nomergebed)
type: basic:file

variant_tables

label: Variant tabels (snpEff)
type: list:basic:file

Amplicon table¶

data:varianttable:ampliconamplicon-table (data:masterfile:amplicon master_file, data:coverage coverage, list:data:snpeff annot_vars, basic:boolean all_amplicons, basic:string table_name)[Source: v1.2.1]

Create variant table for use together with the genome browser.

master_file

label: Master file
type: data:masterfile:amplicon

coverage

label: Amplicon coverage
type: data:coverage

annot_vars

label: Annotated variants
type: list:data:snpeff

all_amplicons

label: Report all amplicons
type: basic:boolean
default: False

table_name

label: Amplicon table name
type: basic:string
default: Amplicons containing variants

variant_table

label: Variant table
type: basic:json

Annotate novel splice junctions (regtools)¶

data:junctions:regtoolsregtools-junctions-annotate (data:seq:nucleotide genome, data:annotation:gtf annotation, data:alignment:bam:star alignment_star, data:alignment:bam alignment, data:bed input_bed_junctions)[Source: v1.1.1]

Identify novel splice junctions by using regtools to annotate against a reference. The process accepts reference genome, reference genome annotation (GTF), and input with reads information (STAR aligment or reads aligned by any other aligner or junctions in BED12 format). If STAR aligner data is given as input, the process calculates BED12 file from STAR ‘SJ.out.tab’ file, and annotates all junctions with ‘regtools junctions annotate’ command. When reads are aligned by other aligner, junctions are extracted with ‘regtools junctions extract’ tool and then annotated with ‘junction annotate’ command. Third option allows user to provide directly BED12 file with junctions, which are then annotated. Finnally, annotated novel junctions are filtered in a separate output file. More information can be found in the [regtools manual](https://regtools.readthedocs.io/en/latest/).

genome

label: Reference genome
type: data:seq:nucleotide

annotation

label: Reference genome annotation (GTF)
type: data:annotation:gtf

alignment_star

label: STAR alignment
type: data:alignment:bam:star
description: Splice junctions detected by STAR aligner (SJ.out.tab STAR output file). Please provide one input ‘STAR alignment’ or ‘Alignment’ by any aligner or directly ‘Junctions in BED12 format’.
required: False

alignment

label: Alignment
type: data:alignment:bam
description: Aligned reads from which splice junctions are going to be extracted. Please provide one input ‘STAR alignment’ or ‘Alignment’ by any aligner or directly ‘Junctions in BED12 format’.
required: False

input_bed_junctions

label: Junctions in BED12 format
type: data:bed
description: Splice junctions in BED12 format. Please provide one input ‘STAR alignment’ or ‘Alignment’ by any aligner or directly ‘Junctions in BED12 format’.
required: False

novel_splice_junctions

label: Table of annotated novel splice junctions
type: basic:file

splice_junctions

label: Table of annotated splice junctions
type: basic:file

novel_sj_bed

label: Novel splice junctions in BED format
type: basic:file

bed

label: Splice junctions in BED format
type: basic:file

novel_sj_bigbed_igv_ucsc

label: Novel splice junctions in BigBed format
type: basic:file
required: False

bigbed_igv_ucsc

label: Splice junctions in BigBed format
type: basic:file
required: False

novel_sj_tbi_jbrowse

label: Novel splice junctions bed tbi index for JBrowse
type: basic:file

tbi_jbrowse

label: Bed tbi index for JBrowse
type: basic:file

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Archive and make multi-sample report for amplicon data¶

data:archive:samples:ampliconamplicon-archive-multi-report (list:data data, list:basic:string fields, basic:boolean j)[Source: v0.3.1]

Create an archive of output files. The ouput folder structure is organized by sample slug and data object’s output-field names. Additionally, create multi-sample report for selected samples.

data

label: Data list
type: list:data

fields

label: Output file fields
type: list:basic:string

j

label: Junk paths
type: basic:boolean
description: Store just names of saved files (junk the path)
default: False

archive

label: Archive of selected samples and a heatmap comparing them
type: basic:file

Archive samples¶

data:archive:samplesarchive-samples (list:data data, list:basic:string fields, basic:boolean j)[Source: v0.4.1]

Create an archive of output files. The ouput folder structure is organized by sample slug and data object’s output-field names.

data

label: Data list
type: list:data

fields

label: Output file fields
type: list:basic:string

j

label: Junk paths
type: basic:boolean
description: Store just names of saved files (junk the path)
default: False

archive

label: Archive
type: basic:file

BAM file¶

data:alignment:bam:uploadupload-bam (basic:file src, basic:string species, basic:string build)[Source: v1.6.1]

Import a BAM file (.bam), which is the binary format for storing sequence alignment data. This format is described on the [SAM Tools web site](http://samtools.github.io/hts-specs/).

src

label: Mapping (BAM)
type: basic:file
description: A mapping file in BAM format. The file will be indexed on upload, so additional BAI files are not required.
validate_regex: \.(bam)$

species

label

Species

type

basic:string

description

Species latin name.

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Build
type: basic:string

bam

label: Uploaded file
type: basic:file

bai

label: Index BAI
type: basic:file

stats

label: Alignment statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BAM file and index¶

data:alignment:bam:uploadupload-bam-indexed (basic:file src, basic:file src2, basic:string species, basic:string build)[Source: v1.6.1]

Import a BAM file (.bam) and BAM index (.bam.bai). BAM file is the binary format for storing sequence alignment data. This format is described on the [SAM Tools web site](http://samtools.github.io/hts-specs/).

src

label: Mapping (BAM)
type: basic:file
description: A mapping file in BAM format.
validate_regex: \.(bam)$

src2

label: bam index (*.bam.bai file)
type: basic:file
description: An index file of a BAM mapping file (ending with bam.bai).
validate_regex: \.(bam.bai)$

species

label

Species

type

basic:string

description

Species latin name.

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Build
type: basic:string

bam

label: Uploaded file
type: basic:file

bai

label: Index BAI
type: basic:file

stats

label: Alignment statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BBDuk (paired-end)¶

data:reads:fastq:paired:bbdukbbduk-paired (data:reads:fastq:paired reads, basic:integer min_length, basic:boolean show_advanced, list:data:seq:nucleotide sequences, list:basic:string literal_sequences, basic:integer kmer_length, basic:boolean check_reverse_complements, basic:boolean mask_middle_base, basic:integer min_kmer_hits, basic:decimal min_kmer_fraction, basic:decimal min_coverage_fraction, basic:integer hamming_distance, basic:integer query_hamming_distance, basic:integer edit_distance, basic:integer hamming_distance2, basic:integer query_hamming_distance2, basic:integer edit_distance2, basic:boolean forbid_N, basic:boolean remove_if_either_bad, basic:boolean find_best_match, basic:boolean perform_error_correction, basic:string k_trim, basic:string k_mask, basic:boolean mask_fully_covered, basic:integer min_k, basic:string quality_trim, basic:integer trim_quality, basic:integer trim_poly_A, basic:decimal min_length_fraction, basic:integer max_length, basic:integer min_average_quality, basic:integer min_average_quality_bases, basic:integer min_base_quality, basic:integer min_consecutive_bases, basic:integer trim_pad, basic:boolean trim_by_overlap, basic:boolean strict_overlap, basic:integer min_overlap, basic:integer min_insert, basic:boolean trim_pairs_evenly, basic:integer force_trim_left, basic:integer force_trim_right, basic:integer force_trim_right2, basic:integer force_trim_mod, basic:integer restrict_left, basic:integer restrict_right, basic:decimal min_GC, basic:decimal max_GC, basic:integer maxns, basic:boolean toss_junk, basic:boolean chastity_filter, basic:boolean barcode_filter, list:data:seq:nucleotide barcode_files, list:basic:string barcode_sequences, basic:integer x_min, basic:integer y_min, basic:integer x_max, basic:integer y_max, basic:decimal entropy, basic:integer entropy_window, basic:integer entropy_k, basic:boolean entropy_mask, basic:integer min_base_frequency, basic:boolean nogroup)[Source: v2.4.1]

BBDuk combines the most common data-quality-related trimming, filtering, and masking operations into a single high-performance tool. It is capable of quality-trimming and filtering, adapter-trimming, contaminant-filtering via kmer matching, sequence masking, GC-filtering, length filtering, entropy-filtering, format conversion, histogram generation, subsampling, quality-score recalibration, kmer cardinality estimation, and various other operations in a single pass. See [here](https://jgi.doe.gov/data-and-tools/bbtools/bb-tools-user-guide/bbduk-guide/) for more information.

reads

label: Reads
type: data:reads:fastq:paired

min_length

label: Minimum length [minlength=10]
type: basic:integer
description: Reads shorter than the minimum length will be discarded after trimming.
default: 10

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

reference.sequences

label: Sequences [ref]
type: list:data:seq:nucleotide
description: Reference sequences include adapters, contaminants, and degenerate sequences. They can be provided in a multi-sequence FASTA file or as a set of literal sequences below.
required: False

reference.literal_sequences

label: Literal sequences [literal]
type: list:basic:string
description: Literal sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

processing.kmer_length

label: Kmer length [k=27]
type: basic:integer
description: Kmer length used for finding contaminants. Contaminants shorter than kmer length will not be found. Kmer length must be at least 1.
default: 27

processing.check_reverse_complements

label: Look for reverse complements of kmers in addition to forward kmers [rcomp=t]
type: basic:boolean
default: True

processing.mask_middle_base

label: Treat the middle base of a kmer as a wildcard to increase sensitivity in the presence of errors [maskmiddle=t]
type: basic:boolean
default: True

processing.min_kmer_hits

label: Minimum number of kmer hits [minkmerhits=1]
type: basic:integer
description: Reads need at least this many matching kmers to be considered as matching the reference.
default: 1

processing.min_kmer_fraction

label: Minimum kmer fraction [minkmerfraction=0.0]
type: basic:decimal
description: A read needs at least this fraction of its total kmers to hit a reference in order to be considered a match. If this and ‘Minimum number of kmer hits’ are set, the greater is used.
default: 0.0

processing.min_coverage_fraction

label: Minimum coverage fraction [mincovfraction=0.0]
type: basic:decimal
description: A read needs at least this fraction of its total bases to be covered by reference kmers to be considered a match. If specified, ‘Minimum coverage fraction’ overrides ‘Minimum number of kmer hits’ and ‘Minimum kmer fraction’.
default: 0.0

processing.hamming_distance

label: Maximum Hamming distance for kmers (substitutions only) [hammingdistance=0]
type: basic:integer
default: 0

processing.query_hamming_distance

label: Hamming distance for query kmers [qhdist=0]
type: basic:integer
default: 0

processing.edit_distance

label: Maximum edit distance from reference kmers (substitutions and indels) [editdistance=0]
type: basic:integer
default: 0

processing.hamming_distance2

label: Hamming distance for short kmers when looking for shorter kmers [hammingdistance2=0]
type: basic:integer
default: 0

processing.query_hamming_distance2

label: Hamming distance for short query kmers when looking for shorter kmers [qhdist2=0]
type: basic:integer
default: 0

processing.edit_distance2

label: Maximum edit distance from short reference kmers (substitutions and indels) when looking for shorter kmers [editdistance2=0]
type: basic:integer
default: 0

processing.forbid_N

label: Forbid matching of read kmers containing N [forbidn=f]
type: basic:boolean
description: By default, these will match a reference ‘A’ if ‘Maximum Hamming distance for kmers’ > 0 or ‘Maximum edit distance from reference kmers’ > 0, to increase sensitivity.
default: False

processing.remove_if_either_bad

label: Remove both sequences of a paired-end read, if either of them is to be removed [removeifeitherbad=t]
type: basic:boolean
default: True

processing.find_best_match

label: If multiple matches, associate read with sequence sharing most kmers [findbestmatch=t]
type: basic:boolean
default: True

processing.perform_error_correction

label: Perform error correction with BBMerge prior to kmer operations [ecco=f]
type: basic:boolean
default: False

operations.k_trim

label

Trimming protocol to remove bases matching reference kmers from reads [ktrim=f]

type

basic:string

default

f

choices

Don’t trim: f
Trim to the right: r
Trim to the left: l

operations.k_mask

label: Symbol to replace bases matching reference kmers [kmask=f]
type: basic:string
description: Allows any non-whitespace character other than t or f. Processes short kmers on both ends.
default: f

operations.mask_fully_covered

label: Only mask bases that are fully covered by kmers [maskfullycovered=f]
type: basic:boolean
default: False

operations.min_k

label: Look for shorter kmers at read tips down to this length when k-trimming or masking [mink=0]
type: basic:integer
description: -1 means disabled. Enabling this will disable treating the middle base of a kmer as a wildcard to increase sensitivity in the presence of errors.
default: -1

operations.quality_trim

label

Trimming protocol to remove bases with quality below the minimum average region quality from read ends [qtrim=f]

type

basic:string

description

Performed after looking for kmers. If enabled, set also ‘Average quality below which to trim region’.

default

f

choices

Trim neither end: f
Trim both ends: rl
Trim only right end: r
Trim only left end: l
Use sliding window: w

operations.trim_quality

label: Average quality below which to trim region [trimq=6]
type: basic:integer
description: Set trimming protocol to enable this parameter.
disabled: operations.quality_trim == ‘f’
default: 6

operations.trim_poly_A

label: Minimum length of poly-A or poly-T tails to trim on either end of reads [trimpolya=0]
type: basic:integer
default: 0

operations.min_length_fraction

label: Minimum length fraction [mlf=0.0]
type: basic:decimal
description: Reads shorter than this fraction of original length after trimming will be discarded.
default: 0.0

operations.max_length

label: Maximum length [maxlength]
type: basic:integer
description: Reads longer than this after trimming will be discarded.
required: False

operations.min_average_quality

label: Minimum average quality [minavgquality=0]
type: basic:integer
description: Reads with average quality (after trimming) below this will be discarded.
default: 0

operations.min_average_quality_bases

label: Number of initial bases to calculate minimum average quality from [maqb=0]
type: basic:integer
description: Used only if positive.
default: 0

operations.min_base_quality

label: Minimum base quality below which reads are discarded after trimming [minbasequality=0]
type: basic:integer
default: 0

operations.min_consecutive_bases

label: Minimum number of consecutive called bases [mcb=0]
type: basic:integer
default: 0

operations.trim_pad

label: Number of bases to trim around matching kmers [tp=0]
type: basic:integer
default: 0

operations.trim_by_overlap

label: Trim adapters based on where paired-end reads overlap [tbo=f]
type: basic:boolean
default: False

operations.strict_overlap

label: Adjust sensitivity in ‘Trim adapters based on where paired-end reads overlap’ mode [strictoverlap=t]
type: basic:boolean
default: True

operations.min_overlap

label: Minimum number of overlapping bases [minoverlap=14]
type: basic:integer
description: Require this many bases of overlap for detection.
default: 14

operations.min_insert

label: Minimum insert size [mininsert=40]
type: basic:integer
description: Require insert size of at least this for overlap. Should be reduced to 16 for small RNA sequencing.
default: 40

operations.trim_pairs_evenly

label: Trim both sequences of paired-end reads to the minimum length of either sequence [tpe=f]
type: basic:boolean
default: False

operations.force_trim_left

label: Position from which to trim bases to the left [forcetrimleft=0]
type: basic:integer
default: 0

operations.force_trim_right

label: Position from which to trim bases to the right [forcetrimright=0]
type: basic:integer
default: 0

operations.force_trim_right2

label: Number of bases to trim from the right end [forcetrimright2=0]
type: basic:integer
default: 0

operations.force_trim_mod

label: Modulo to right-trim reads [forcetrimmod=0]
type: basic:integer
description: Trim reads to the largest multiple of modulo.
default: 0

operations.restrict_left

label: Number of leftmost bases to look in for kmer matches [restrictleft=0]
type: basic:integer
default: 0

operations.restrict_right

label: Number of rightmosot bases to look in for kmer matches [restrictright=0]
type: basic:integer
default: 0

operations.min_GC

label: Minimum GC content [mingc=0.0]
type: basic:decimal
description: Discard reads with lower GC content.
default: 0.0

operations.max_GC

label: Maximum GC content [maxgc=1.0]
type: basic:decimal
description: Discard reads with higher GC content.
default: 1.0

operations.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
default: -1

operations.toss_junk

label: Discard reads with invalid characters as bases [tossjunk=f]
type: basic:boolean
default: False

header_parsing.chastity_filter

label: Discard reads that fail Illumina chastity filtering [chastityfilter=f]
type: basic:boolean
description: Discard reads with id containing ‘ 1:Y:’ or ‘ 2:Y:’.
default: False

header_parsing.barcode_filter

label: Remove reads with unexpected barcodes if barcodes are set, or barcodes containing ‘N’ otherwise [barcodefilter=f]
type: basic:boolean
description: A barcode must be the last part of the read header.
default: False

header_parsing.barcode_files

label: Barcode sequences [barcodes]
type: list:data:seq:nucleotide
required: False

header_parsing.barcode_sequences

label: Literal barcode sequences [barcodes]
type: list:basic:string
description: Literal barcode sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

header_parsing.x_min

label: Minimum X coordinate [xmin=-1]
type: basic:integer
description: If positive, discard reads with a smaller X coordinate.
default: -1

header_parsing.y_min

label: Minimum Y coordinate [ymin=-1]
type: basic:integer
description: If positive, discard reads with a smaller Y coordinate.
default: -1

header_parsing.x_max

label: Maximum X coordinate [xmax=-1]
type: basic:integer
description: If positive, discard reads with a larger X coordinate.
default: -1

header_parsing.y_max

label: Maximum Y coordinate [ymax=-1]
type: basic:integer
description: If positive, discard reads with a larger Y coordinate.
default: -1

complexity.entropy

label: Minimum entropy [entropy=-1.0]
type: basic:decimal
description: Set between 0 and 1 to filter reads with entropy below that value. Higher is more stringent.
default: -1.0

complexity.entropy_window

label: Length of sliding window used to calculate entropy [entropywindow=50]
type: basic:integer
description: To use the sliding window set minimum entropy in range between 0.0 and 1.0.
default: 50

complexity.entropy_k

label: Length of kmers used to calcuate entropy [entropyk=5]
type: basic:integer
default: 5

complexity.entropy_mask

label: Mask low-entropy parts of sequences with N instead of discarding [entropymask=f]
type: basic:boolean
default: False

complexity.min_base_frequency

label: Minimum base frequency [minbasefrequency=0]
type: basic:integer
default: 0

fastqc.nogroup

label: Disable grouping of bases for reads >50bp [nogroup]
type: basic:boolean
description: All reports will show data for every base in the read. Using this option will cause fastqc to crash and burn if you use it on really long reads.
default: False

fastq

label: Remaining upstream reads
type: list:basic:file

fastq2

label: Remaining downstream reads
type: list:basic:file

statistics

label: Statistics
type: list:basic:file

fastqc_url

label: Upstream quality control with FastQC
type: list:basic:file:html

fastqc_url2

label: Downstream quality control with FastQC
type: list:basic:file:html

fastqc_archive

label: Download upstream FastQC archive
type: list:basic:file

fastqc_archive2

label: Download downstream FastQC archive
type: list:basic:file

BBDuk (single-end)¶

data:reads:fastq:single:bbdukbbduk-single (data:reads:fastq:single reads, basic:integer min_length, basic:boolean show_advanced, list:data:seq:nucleotide sequences, list:basic:string literal_sequences, basic:integer kmer_length, basic:boolean check_reverse_complements, basic:boolean mask_middle_base, basic:integer min_kmer_hits, basic:decimal min_kmer_fraction, basic:decimal min_coverage_fraction, basic:integer hamming_distance, basic:integer query_hamming_distance, basic:integer edit_distance, basic:integer hamming_distance2, basic:integer query_hamming_distance2, basic:integer edit_distance2, basic:boolean forbid_N, basic:boolean find_best_match, basic:string k_trim, basic:string k_mask, basic:boolean mask_fully_covered, basic:integer min_k, basic:string quality_trim, basic:integer trim_quality, basic:integer trim_poly_A, basic:decimal min_length_fraction, basic:integer max_length, basic:integer min_average_quality, basic:integer min_average_quality_bases, basic:integer min_base_quality, basic:integer min_consecutive_bases, basic:integer trim_pad, basic:integer min_overlap, basic:integer min_insert, basic:integer force_trim_left, basic:integer force_trim_right, basic:integer force_trim_right2, basic:integer force_trim_mod, basic:integer restrict_left, basic:integer restrict_right, basic:decimal min_GC, basic:decimal max_GC, basic:integer maxns, basic:boolean toss_junk, basic:boolean chastity_filter, basic:boolean barcode_filter, list:data:seq:nucleotide barcode_files, list:basic:string barcode_sequences, basic:integer x_min, basic:integer y_min, basic:integer x_max, basic:integer y_max, basic:decimal entropy, basic:integer entropy_window, basic:integer entropy_k, basic:boolean entropy_mask, basic:integer min_base_frequency, basic:boolean nogroup)[Source: v2.4.1]

BBDuk combines the most common data-quality-related trimming, filtering, and masking operations into a single high-performance tool. It is capable of quality-trimming and filtering, adapter-trimming, contaminant-filtering via kmer matching, sequence masking, GC-filtering, length filtering, entropy-filtering, format conversion, histogram generation, subsampling, quality-score recalibration, kmer cardinality estimation, and various other operations in a single pass. See [here](https://jgi.doe.gov/data-and-tools/bbtools/bb-tools-user-guide/bbduk-guide/) for more information.

reads

label: Reads
type: data:reads:fastq:single

min_length

label: Minimum length [minlength=10]
type: basic:integer
description: Reads shorter than the minimum length will be discarded after trimming.
default: 10

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

reference.sequences

label: Sequences [ref]
type: list:data:seq:nucleotide
description: Reference sequences include adapters, contaminants, and degenerate sequences. They can be provided in a multi-sequence FASTA file or as a set of literal sequences below.
required: False

reference.literal_sequences

label: Literal sequences [literal]
type: list:basic:string
description: Literal sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

processing.kmer_length

label: Kmer length [k=27]
type: basic:integer
description: Kmer length used for finding contaminants. Contaminants shorter than Kmer length will not be found. Kmer length must be at least 1.
default: 27

processing.check_reverse_complements

label: Look for reverse complements of kmers in addition to forward kmers [rcomp=t]
type: basic:boolean
default: True

processing.mask_middle_base

label: Treat the middle base of a kmer as a wildcard to increase sensitivity in the presence of errors [maskmiddle=t]
type: basic:boolean
default: True

processing.min_kmer_hits

label: Minimum number of kmer hits [minkmerhits=1]
type: basic:integer
description: Reads need at least this many matching kmers to be considered matching the reference.
default: 1

processing.min_kmer_fraction

label: Minimum kmer fraction [minkmerfraction=0.0]
type: basic:decimal
description: A read needs at least this fraction of its total kmers to hit a reference in order to be considered a match. If this and ‘Minimum number of kmer hits’ are set, the greater is used.
default: 0.0

processing.min_coverage_fraction

label: Minimum coverage fraction [mincovfraction=0.0]
type: basic:decimal
description: A read needs at least this fraction of its total bases to be covered by reference kmers to be considered a match. If specified, ‘Minimum coverage fraction’ overrides ‘Minimum number of kmer hits’ and ‘Minimum kmer fraction’.
default: 0.0

processing.hamming_distance

label: Maximum Hamming distance for kmers (substitutions only) [hammingdistance=0]
type: basic:integer
default: 0

processing.query_hamming_distance

label: Hamming distance for query kmers [qhdist=0]
type: basic:integer
default: 0

processing.edit_distance

label: Maximum edit distance from reference kmers (substitutions and indels) [editdistance=0]
type: basic:integer
default: 0

processing.hamming_distance2

label: Hamming distance for short kmers when looking for shorter kmers [hammingdistance2=0]
type: basic:integer
default: 0

processing.query_hamming_distance2

label: Hamming distance for short query kmers when looking for shorter kmers [qhdist2=0]
type: basic:integer
default: 0

processing.edit_distance2

label: Maximum edit distance from short reference kmers (substitutions and indels) when looking for shorter kmers [editdistance2=0]
type: basic:integer
default: 0

processing.forbid_N

label: Forbid matching of read kmers containing N [forbidn=f]
type: basic:boolean
description: By default, these will match a reference ‘A’ if ‘Maximum Hamming distance for kmers’ > 0 or ‘Maximum edit distance from reference kmers’ > 0, to increase sensitivity.
default: False

processing.find_best_match

label: If multiple matches, associate read with sequence sharing most kmers [findbestmatch=f]
type: basic:boolean
default: True

operations.k_trim

label

Trimming protocol to remove bases matching reference kmers from reads [ktrim=f]

type

basic:string

default

f

choices

Don’t trim: f
Trim to the right: r
Trim to the left: l

operations.k_mask

label: Symbol to replace bases matching reference kmers [kmask=f]
type: basic:string
description: Allows any non-whitespace character other than t or f. Processes short kmers on both ends.
default: f

operations.mask_fully_covered

label: Only mask bases that are fully covered by kmers [maskfullycovered=f]
type: basic:boolean
default: False

operations.min_k

label: Look for shorter kmers at read tips down to this length when k-trimming or masking [mink=0]
type: basic:integer
description: -1 means disabled. Enabling this will disable treating the middle base of a kmer as a wildcard to increase sensitivity in the presence of errors.
default: -1

operations.quality_trim

label

Trimming protocol to remove bases with quality below the minimum average region quality from read ends [qtrim=f]

type

basic:string

description

Performed after looking for kmers. If enabled, set also ‘Average quality below which to trim region’.

default

f

choices

Trim neither end: f
Trim both ends: rl
Trim only right end: r
Trim only left end: l
Use sliding window: w

operations.trim_quality

label: Average quality below which to trim region [trimq=6]
type: basic:integer
description: Set trimming protocol to enable this parameter.
disabled: operations.quality_trim == ‘f’
default: 6

operations.trim_poly_A

label: Minimum length of poly-A or poly-T tails to trim on either end of reads [trimpolya=0]
type: basic:integer
default: 0

operations.min_length_fraction

label: Minimum length fraction [mlf=0]
type: basic:decimal
description: Reads shorter than this fraction of original length after trimming will be discarded.
default: 0.0

operations.max_length

label: Maximum length [maxlength]
type: basic:integer
description: Reads longer than this after trimming will be discarded.
required: False

operations.min_average_quality

label: Minimum average quality [minavgquality=0]
type: basic:integer
description: Reads with average quality (after trimming) below this will be discarded.
default: 0

operations.min_average_quality_bases

label: Number of initial bases to calculate minimum average quality from [maqb=0]
type: basic:integer
description: Used only if positive.
default: 0

operations.min_base_quality

label: Minimum base quality below which reads are discarded after trimming [minbasequality=0]
type: basic:integer
default: 0

operations.min_consecutive_bases

label: Minimum number of consecutive called bases [mcb=0]
type: basic:integer
default: 0

operations.trim_pad

label: Number of bases to trim around matching kmers [tp=0]
type: basic:integer
default: 0

operations.min_overlap

label: Minimum number of overlapping bases [minoverlap=14]
type: basic:integer
description: Require this many bases of overlap for detection.
default: 14

operations.min_insert

label: Minimum insert size [mininsert=40]
type: basic:integer
description: Require insert size of at least this for overlap. Should be reduced to 16 for small RNA sequencing.
default: 40

operations.force_trim_left

label: Position from which to trim bases to the left [forcetrimleft=0]
type: basic:integer
default: 0

operations.force_trim_right

label: Position from which to trim bases to the right [forcetrimright=0]
type: basic:integer
default: 0

operations.force_trim_right2

label: Number of bases to trim from the right end [forcetrimright2=0]
type: basic:integer
default: 0

operations.force_trim_mod

label: Modulo to right-trim reads [forcetrimmod=0]
type: basic:integer
description: Trim reads to the largest multiple of modulo.
default: 0

operations.restrict_left

label: Number of leftmost bases to look in for kmer matches [restrictleft=0]
type: basic:integer
default: 0

operations.restrict_right

label: Number of rightmosot bases to look in for kmer matches [restricright=0]
type: basic:integer
default: 0

operations.min_GC

label: Minimum GC content [mingc=0.0]
type: basic:decimal
description: Discard reads with lower GC content.
default: 0.0

operations.max_GC

label: Maximum GC content [maxgc=1.0]
type: basic:decimal
description: Discard reads with higher GC content.
default: 1.0

operations.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
default: -1

operations.toss_junk

label: Discard reads with invalid characters as bases [tossjunk=f]
type: basic:boolean
default: False

header_parsing.chastity_filter

label: Discard reads that fail Illumina chastity filtering [chastityfilter=f]
type: basic:boolean
description: Discard reads with id containing ‘ 1:Y:’ or ‘ 2:Y:’.
default: False

header_parsing.barcode_filter

label: Remove reads with unexpected barcodes if barcodes are set, or barcodes containing ‘N’ otherwise [barcodefilter=f]
type: basic:boolean
description: A barcode must be the last part of the read header.
default: False

header_parsing.barcode_files

label: Barcode sequences [barcodes]
type: list:data:seq:nucleotide
required: False

header_parsing.barcode_sequences

label: Literal barcode sequences [barcodes]
type: list:basic:string
description: Literal barcode sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

header_parsing.x_min

label: Minimum X coordinate [xmin=-1]
type: basic:integer
description: If positive, discard reads with a smaller X coordinate.
default: -1

header_parsing.y_min

label: Minimum Y coordinate [ymin=-1]
type: basic:integer
description: If positive, discard reads with a smaller Y coordinate.
default: -1

header_parsing.x_max

label: Maximum X coordinate [xmax=-1]
type: basic:integer
description: If positive, discard reads with a larger X coordinate.
default: -1

header_parsing.y_max

label: Maximum Y coordinate [ymax=-1]
type: basic:integer
description: If positive, discard reads with a larger Y coordinate.
default: -1

complexity.entropy

label: Minimum entropy [entropy=-1]
type: basic:decimal
description: Set between 0 and 1 to filter reads with entropy below that value. Higher is more stringent.
default: -1.0

complexity.entropy_window

label: Length of sliding window used to calculate entropy [entropywindow=50]
type: basic:integer
description: To use the sliding window set minimum entropy in range between 0.0 and 1.0.
default: 50

complexity.entropy_k

label: Length of kmers used to calcuate entropy [entropyk=5]
type: basic:integer
default: 5

complexity.entropy_mask

label: Mask low-entropy parts of sequences with N instead of discarding [entropymask=f]
type: basic:boolean
default: False

complexity.min_base_frequency

label: Minimum base frequency [minbasefrequency=0]
type: basic:integer
default: 0

fastqc.nogroup

label: Disable grouping of bases for reads >50bp [nogroup]
type: basic:boolean
description: All reports will show data for every base in the read. Using this option will cause fastqc to crash and burn if you use it on really long reads.
default: False

fastq

label: Remaining reads
type: list:basic:file

statistics

label: Statistics
type: list:basic:file

fastqc_url

label: Quality control with FastQC
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive
type: list:basic:file

BBDuk - STAR - FeatureCounts (3’ mRNA-Seq, paired-end)¶

data:workflow:quant:featurecounts:pairedworkflow-bbduk-star-fc-quant-paired (data:reads:fastq:paired reads, data:index:star star_index, list:data:seq:nucleotide adapters, data:annotation annotation, basic:string stranded, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, data:index:star rrna_reference, data:index:star globin_reference)[Source: v2.0.1]

This 3’ mRNA-Seq pipeline is comprised of QC, preprocessing, alignment and quantification steps. Reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Preprocessed reads are aligned by __STAR__ aligner. For read-count quantification, the __FeatureCounts__ tool is used. QC steps include downsampling, QoRTs QC analysis and alignment of input reads to the rRNA/globin reference sequences. The reported alignment rate is used to asses the rRNA/globin sequence depletion rate.

reads

label: Paired-end reads
type: data:reads:fastq:paired

star_index

label: Star index
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

adapters

label: Adapters
type: list:data:seq:nucleotide
description: Provide a list of sequencing adapters files (.fasta) to be removed by BBDuk.
required: False

annotation

label: Annotation
type: data:annotation

stranded

label

Select the type of kit used for library preparation.

type

basic:string

choices

Strand-specific forward: forward
Strand-specific reverse: reverse

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.advanced.seed

label: Seed
type: basic:integer
default: 11

downsampling.advanced.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads in range [0.0, 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.advanced.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

qc.rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

qc.globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

BBDuk - STAR - FeatureCounts (3’ mRNA-Seq, single-end)¶

data:workflow:quant:featurecounts:singleworkflow-bbduk-star-fc-quant-single (data:reads:fastq:single reads, data:index:star star_index, list:data:seq:nucleotide adapters, data:annotation annotation, basic:string stranded, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, data:index:star rrna_reference, data:index:star globin_reference)[Source: v2.0.1]

This 3’ mRNA-Seq pipeline is comprised of QC, preprocessing, alignment and quantification steps. Reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Preprocessed reads are aligned by __STAR__ aligner. For read-count quantification, the __FeatureCounts__ tool is used. QC steps include downsampling, QoRTs QC analysis and alignment of input reads to the rRNA/globin reference sequences. The reported alignment rate is used to asses the rRNA/globin sequence depletion rate.

reads

label: Input single-end reads
type: data:reads:fastq:single

star_index

label: Star index
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

adapters

label: Adapters
type: list:data:seq:nucleotide
description: Provide a list of sequencing adapters files (.fasta) to be removed by BBDuk.
required: False

annotation

label: Annotation
type: data:annotation

stranded

label

Select the type of kit used for library preparation.

type

basic:string

choices

Strand-specific forward: forward
Strand-specific reverse: reverse

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.advanced.seed

label: Seed
type: basic:integer
default: 11

downsampling.advanced.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads in range [0.0, 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.advanced.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

qc.rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

qc.globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

BBDuk - STAR - HTSeq-count (paired-end)¶

data:workflow:rnaseq:htseq:pairedworkflow-bbduk-star-htseq-paired (data:reads:fastq:paired reads, data:index:star star_index, list:data:seq:nucleotide adapters, data:annotation annotation, basic:string stranded)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Compared to similar tools, BBDuk is regarded for its computational efficiency. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.nature.com/articles/nmeth.4106). Finally, aligned reads are summarized to genes by __HTSeq-count__. Compared to featureCounts, HTSeq-count is not as computationally efficient. All three tools in this workflow support parallelization to accelerate the analysis.

reads

label: Paired-end reads
type: data:reads:fastq:paired

star_index

label: Star index
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

adapters

label: Adapters
type: list:data:seq:nucleotide
description: Provide a list of sequencing adapters files (.fasta) to be removed by BBDuk.
required: False

annotation

label: Annotation
type: data:annotation

stranded

label

Select the QuantSeq kit used for library preparation.

type

basic:string

choices

QuantSeq FWD: yes
QuantSeq REV: reverse

BBDuk - STAR - HTSeq-count (single-end)¶

data:workflow:rnaseq:htseq:singleworkflow-bbduk-star-htseq (data:reads:fastq:single reads, data:index:star star_index, list:data:seq:nucleotide adapters, data:annotation annotation, basic:string stranded)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Compared to similar tools, BBDuk is regarded for its computational efficiency. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.nature.com/articles/nmeth.4106). Finally, aligned reads are summarized to genes by __HTSeq-count__. Compared to featureCounts, HTSeq-count is not as computationally efficient. All three tools in this workflow support parallelization to accelerate the analysis.

reads

label: Input single-end reads
type: data:reads:fastq:single

star_index

label: Star index
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

adapters

label: Adapters
type: list:data:seq:nucleotide
description: Provide a list of sequencing adapters files (.fasta) to be removed by BBDuk.
required: False

annotation

label: annotation
type: data:annotation

stranded

label

Select the QuantSeq kit used for library preparation.

type

basic:string

choices

QuantSeq FWD: yes
QuantSeq REV: reverse

BBDuk - STAR - featureCounts - QC (paired-end)¶

data:workflow:rnaseq:featurecounts:qcworkflow-bbduk-star-featurecounts-qc-paired (data:reads:fastq:paired reads, list:data:seq:nucleotide adapters, basic:boolean show_advanced, list:basic:string custom_adapter_sequences, basic:integer kmer_length, basic:integer min_k, basic:integer hamming_distance, basic:integer maxns, basic:integer trim_quality, basic:integer min_length, data:index:star genome, basic:boolean show_advanced, basic:boolean unstranded, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer outFilterScoreMin, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax, basic:string alignEndsType, basic:string outSAMunmapped, basic:string outSAMattributes, basic:string outSAMattrRGline, data:annotation annotation, basic:boolean show_advanced, basic:string assay_type, data:index:salmon cdna_index, basic:integer n_reads, basic:string feature_class, basic:string feature_type, basic:string id_attribute, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, data:index:star rrna_reference, data:index:star globin_reference)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps preprocessing, alignment, and quantification. First, reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Compared to similar tools, BBDuk is regarded for its computational efficiency. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5792058/). Finally, aligned reads are summarized to genes by __featureCounts__. Gaining wide adoption among the bioinformatics community, featureCounts yields expressions in a computationally efficient manner. All three tools in this workflow support parallelization to accelerate the analysis. rRNA contamination rate in the sample is determined using the STAR aligner. Quality-trimmed reads are down-sampled (using Seqtk tool) and aligned to the rRNA reference sequences. The alignment rate indicates the percentage of the reads in the sample that are derived from the rRNA sequences.

preprocessing.reads

label: Reads
type: data:reads:fastq:paired

preprocessing.adapters

label: Adapters
type: list:data:seq:nucleotide
required: False

preprocessing.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

preprocessing.custom_adapter_sequences

label: Custom adapter sequences [literal]
type: list:basic:string
description: Custom adapter sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
hidden: !preprocessing.show_advanced
default: []

preprocessing.kmer_length

label: K-mer length
type: basic:integer
description: K-mer length must be smaller or equal to the length of adapters.
hidden: !preprocessing.show_advanced
default: 23

preprocessing.min_k

label: Minimum k-mer length at right end of reads used for trimming
type: basic:integer
disabled: preprocessing.adapters.length === 0 && preprocessing.custom_adapter_sequences.length === 0
hidden: !preprocessing.show_advanced
default: 11

preprocessing.hamming_distance

label: Maximum Hamming distance for k-mers
type: basic:integer
hidden: !preprocessing.show_advanced
default: 1

preprocessing.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
hidden: !preprocessing.show_advanced
default: -1

preprocessing.trim_quality

label: Quality below which to trim reads from the right end
type: basic:integer
description: Phred algorithm is used, which is more accurate than naive trimming.
hidden: !preprocessing.show_advanced
default: 10

preprocessing.min_length

label: Minimum read length
type: basic:integer
description: Reads shorter than minimum read length after trimming are discarded.
hidden: !preprocessing.show_advanced
default: 20

alignment.genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

alignment.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

alignment.unstranded

label: The data is unstranded
type: basic:boolean
description: For unstranded RNA-seq data, Cufflinks/Cuffdiff require spliced alignments with XS strand attribute, which STAR will generate with –outSAMstrandField intronMotif option. As required, the XS strand attribute will be generated for all alignments that contain splice junctions. The spliced alignments that have undefined strand (i.e. containing only non-canonical unannotated junctions) will be suppressed. If you have stranded RNA-seq data, you do not need to use any specific STAR options. Instead, you need to run Cufflinks with the library option –library-type options. For example, c ufflinks –library-type fr-firststrand should be used for the standard dUTP protocol, including Illumina’s stranded Tru-Seq. This option has to be used only for Cufflinks runs and not for STAR runs.
hidden: !alignment.show_advanced
default: False

alignment.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
hidden: !alignment.show_advanced
default: False

alignment.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

alignment.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: detect_chimeric.chimeric != true
default: 20

alignment.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: False

alignment.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: t_coordinates.quantmode != true
default: False

alignment.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: t_coordinates.quantmode != true
default: False

alignment.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

alignment.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

alignment.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

alignment.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

alignment.filtering.outFilterScoreMin

label: –outFilterScoreMin
type: basic:integer
description: Alignment will be output only if its score is higher than or equal to this value (default: 0).
required: False

alignment.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

alignment.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

alignment.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

alignment.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

alignment.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

alignment.alignment.alignEndsType

label

–alignEndsType

type

basic:string

description

Type of read ends alignment (default: Local).

required

False

default

Local

choices

Local: Local
EndToEnd: EndToEnd
Extend5pOfRead1: Extend5pOfRead1
Extend5pOfReads12: Extend5pOfReads12

alignment.output_sam_bam.outSAMunmapped

label

–outSAMunmapped

type

basic:string

description

Output of unmapped reads in the SAM format.

required

False

default

None

choices

None: None
Within: Within

alignment.output_sam_bam.outSAMattributes

label

–outSAMattributes

type

basic:string

description

a string of desired SAM attributes, in the order desired for the output SAM.

required

False

default

Standard

choices

None: None
Standard: Standard
All: All

alignment.output_sam_bam.outSAMattrRGline

label: –outSAMattrRGline
type: basic:string
description: SAM/BAM read group line. The first word contains the read group identifier and must start with “ID:”, e.g. –outSAMattrRGline ID:xxx CN:yy “DS:z z z”
required: False

quantification.annotation

label: Annotation
type: data:annotation

quantification.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

quantification.assay_type

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

hidden

!quantification.show_advanced

default

non_specific

choices

Strand non-specific: non_specific
Strand-specific forward: forward
Strand-specific reverse: reverse
Detect automatically: auto

quantification.cdna_index

label: cDNA index file
type: data:index:salmon
description: Transcriptome index file created using the Salmon indexing tool. cDNA (transcriptome) sequences used for index file creation must be derived from the same species as the input sequencing reads to obtain the reliable analysis results.
required: False
hidden: quantification.assay_type != ‘auto’

quantification.n_reads

label: Number of reads in subsampled alignment file
type: basic:integer
description: Alignment (.bam) file subsample size. Increase the number of reads to make automatic detection more reliable. Decrease the number of reads to make automatic detection run faster.
hidden: quantification.assay_type != ‘auto’
default: 5000000

quantification.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GTF/GFF3 file) to be used. All other features will be ignored.
hidden: !quantification.show_advanced
default: exon

quantification.feature_type

label

Feature type

type

basic:string

description

The type of feature the quantification program summarizes over (e.g. gene or transcript-level analysis). The value of this parameter needs to be chosen in line with ‘ID attribute’ below.

hidden

!quantification.show_advanced

default

gene

choices

gene: gene
transcript: transcript

quantification.id_attribute

label

ID attribute

type

basic:string

description

GTF/GFF3 attribute to be used as feature ID. Several GTF/GFF3 lines with the same feature ID are considered as parts of the same feature. The feature ID is used to identify the counts in the output table. In GTF files this is usually ‘gene_id’, in GFF3 files this is often ‘ID’, and ‘transcript_id’ is frequently a valid choice for both annotation formats.

hidden

!quantification.show_advanced

default

gene_id

choices

gene_id: gene_id
transcript_id: transcript_id
ID: ID
geneid: geneid

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.advanced.seed

label: Seed
type: basic:integer
default: 11

downsampling.advanced.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads [0 - 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.advanced.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

qc.rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

qc.globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

BBDuk - STAR - featureCounts - QC (single-end)¶

data:workflow:rnaseq:featurecounts:qcworkflow-bbduk-star-featurecounts-qc-single (data:reads:fastq:single reads, list:data:seq:nucleotide adapters, basic:boolean show_advanced, list:basic:string custom_adapter_sequences, basic:integer kmer_length, basic:integer min_k, basic:integer hamming_distance, basic:integer maxns, basic:integer trim_quality, basic:integer min_length, data:index:star genome, basic:boolean show_advanced, basic:boolean unstranded, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer outFilterScoreMin, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax, basic:string alignEndsType, basic:string outSAMunmapped, basic:string outSAMattributes, basic:string outSAMattrRGline, data:annotation annotation, basic:boolean show_advanced, basic:string assay_type, data:index:salmon cdna_index, basic:integer n_reads, basic:string feature_class, basic:string feature_type, basic:string id_attribute, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, data:index:star rrna_reference, data:index:star globin_reference)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps preprocessing, alignment, and quantification. First, reads are preprocessed by __BBDuk__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Compared to similar tools, BBDuk is regarded for its computational efficiency. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5792058/). Finally, aligned reads are summarized to genes by __featureCounts__. Gaining wide adoption among the bioinformatics community, featureCounts yields expressions in a computationally efficient manner. All three tools in this workflow support parallelization to accelerate the analysis. rRNA contamination rate in the sample is determined using the STAR aligner. Quality-trimmed reads are down-sampled (using Seqtk tool) and aligned to the rRNA reference sequences. The alignment rate indicates the percentage of the reads in the sample that are derived from the rRNA sequences.

preprocessing.reads

label: Reads
type: data:reads:fastq:single

preprocessing.adapters

label: Adapters
type: list:data:seq:nucleotide
required: False

preprocessing.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

preprocessing.custom_adapter_sequences

label: Custom adapter sequences [literal]
type: list:basic:string
description: Custom adapter sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
hidden: !preprocessing.show_advanced
default: []

preprocessing.kmer_length

label: K-mer length
type: basic:integer
description: K-mer length must be smaller or equal to the length of adapters.
hidden: !preprocessing.show_advanced
default: 23

preprocessing.min_k

label: Minimum k-mer length at right end of reads used for trimming
type: basic:integer
disabled: preprocessing.adapters.length === 0 && preprocessing.custom_adapter_sequences.length === 0
hidden: !preprocessing.show_advanced
default: 11

preprocessing.hamming_distance

label: Maximum Hamming distance for k-mers
type: basic:integer
hidden: !preprocessing.show_advanced
default: 1

preprocessing.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
hidden: !preprocessing.show_advanced
default: -1

preprocessing.trim_quality

label: Quality below which to trim reads from the right end
type: basic:integer
description: Phred algorithm is used, which is more accurate than naive trimming.
hidden: !preprocessing.show_advanced
default: 10

preprocessing.min_length

label: Minimum read length
type: basic:integer
description: Reads shorter than minimum read length after trimming are discarded.
hidden: !preprocessing.show_advanced
default: 20

alignment.genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

alignment.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

alignment.unstranded

label: The data is unstranded
type: basic:boolean
description: For unstranded RNA-seq data, Cufflinks/Cuffdiff require spliced alignments with XS strand attribute, which STAR will generate with –outSAMstrandField intronMotif option. As required, the XS strand attribute will be generated for all alignments that contain splice junctions. The spliced alignments that have undefined strand (i.e. containing only non-canonical unannotated junctions) will be suppressed. If you have stranded RNA-seq data, you do not need to use any specific STAR options. Instead, you need to run Cufflinks with the library option –library-type options. For example, c ufflinks –library-type fr-firststrand should be used for the standard dUTP protocol, including Illumina’s stranded Tru-Seq. This option has to be used only for Cufflinks runs and not for STAR runs.
hidden: !alignment.show_advanced
default: False

alignment.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
hidden: !alignment.show_advanced
default: False

alignment.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

alignment.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: detect_chimeric.chimeric != true
default: 20

alignment.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: False

alignment.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: t_coordinates.quantmode != true
default: False

alignment.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: t_coordinates.quantmode != true
default: False

alignment.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

alignment.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

alignment.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

alignment.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

alignment.filtering.outFilterScoreMin

label: –outFilterScoreMin
type: basic:integer
description: Alignment will be output only if its score is higher than or equal to this value (default: 0).
required: False

alignment.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

alignment.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

alignment.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

alignment.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

alignment.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

alignment.alignment.alignEndsType

label

–alignEndsType

type

basic:string

description

Type of read ends alignment (default: Local).

required

False

default

Local

choices

Local: Local
EndToEnd: EndToEnd
Extend5pOfRead1: Extend5pOfRead1
Extend5pOfReads12: Extend5pOfReads12

alignment.output_sam_bam.outSAMunmapped

label

–outSAMunmapped

type

basic:string

description

Output of unmapped reads in the SAM format.

required

False

default

None

choices

None: None
Within: Within

alignment.output_sam_bam.outSAMattributes

label

–outSAMattributes

type

basic:string

description

a string of desired SAM attributes, in the order desired for the output SAM.

required

False

default

Standard

choices

None: None
Standard: Standard
All: All

alignment.output_sam_bam.outSAMattrRGline

label: –outSAMattrRGline
type: basic:string
description: SAM/BAM read group line. The first word contains the read group identifier and must start with “ID:”, e.g. –outSAMattrRGline ID:xxx CN:yy “DS:z z z”
required: False

quantification.annotation

label: Annotation
type: data:annotation

quantification.show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

quantification.assay_type

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

hidden

!quantification.show_advanced

default

non_specific

choices

Strand non-specific: non_specific
Strand-specific forward: forward
Strand-specific reverse: reverse
Detect automatically: auto

quantification.cdna_index

label: cDNA index file
type: data:index:salmon
description: Transcriptome index file created using the Salmon indexing tool. cDNA (transcriptome) sequences used for index file creation must be derived from the same species as the input sequencing reads to obtain the reliable analysis results.
required: False
hidden: quantification.assay_type != ‘auto’

quantification.n_reads

label: Number of reads in subsampled alignment file
type: basic:integer
description: Alignment (.bam) file subsample size. Increase the number of reads to make automatic detection more reliable. Decrease the number of reads to make automatic detection run faster.
hidden: quantification.assay_type != ‘auto’
default: 5000000

quantification.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GTF/GFF3 file) to be used. All other features will be ignored.
hidden: !quantification.show_advanced
default: exon

quantification.feature_type

label

Feature type

type

basic:string

description

The type of feature the quantification program summarizes over (e.g. gene or transcript-level analysis). The value of this parameter needs to be chosen in line with ‘ID attribute’ below.

hidden

!quantification.show_advanced

default

gene

choices

gene: gene
transcript: transcript

quantification.id_attribute

label

ID attribute

type

basic:string

description

GTF/GFF3 attribute to be used as feature ID. Several GTF/GFF3 lines with the same feature ID will be considered as parts of the same feature. The feature ID is used to identify the counts in the output table. In GTF files this is usually ‘gene_id’, in GFF3 files this is often ‘ID’, and ‘transcript_id’ is frequently a valid choice for both annotation formats.

hidden

!quantification.show_advanced

default

gene_id

choices

gene_id: gene_id
transcript_id: transcript_id
ID: ID
geneid: geneid

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.advanced.seed

label: Seed
type: basic:integer
default: 11

downsampling.advanced.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads [0 - 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.advanced.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

qc.rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

qc.globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

BBDuk - Salmon - QC (paired-end)¶

data:workflow:rnaseq:salmonworkflow-bbduk-salmon-qc-paired (data:reads:fastq:paired reads, data:index:salmon salmon_index, data:index:star genome, data:annotation annotation, data:index:star rrna_reference, data:index:star globin_reference, basic:boolean show_advanced, list:data:seq:nucleotide adapters, list:basic:string custom_adapter_sequences, basic:integer kmer_length, basic:integer min_k, basic:integer hamming_distance, basic:integer maxns, basic:integer trim_quality, basic:integer min_length, basic:boolean seq_bias, basic:boolean gc_bias, basic:decimal consensus_slack, basic:decimal min_score_fraction, basic:integer range_factorization_bins, basic:integer min_assigned_frag, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass)[Source: v3.0.1]

Alignment-free RNA-seq pipeline. Salmon tool and tximport package are used in quantification step to produce gene-level abundance estimates. rRNA and globin-sequence contamination rate in the sample is determined using STAR aligner. Quality-trimmed reads are down-sampled (using Seqtk tool) and aligned to the genome, rRNA and globin reference sequences. The rRNA and globin-sequence alignment rates indicate the percentage of the reads in the sample that are of rRNA and globin origin, respectively. Alignment of down-sampled data to a whole genome reference sequence is used to produce an alignment file suitable for Samtools and QoRTs QC analysis. Per-sample analysis results and QC data is summarized by the MultiQC tool.

reads

label: Select sample(s)
type: data:reads:fastq:paired

salmon_index

label: Salmon index
type: data:index:salmon

genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation

rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

preprocessing.adapters

label: Adapters
type: list:data:seq:nucleotide
required: False

preprocessing.custom_adapter_sequences

label: Custom adapter sequences [literal]
type: list:basic:string
description: Custom adapter sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

preprocessing.kmer_length

label: K-mer length
type: basic:integer
description: K-mer length must be smaller or equal to the length of adapters.
default: 23

preprocessing.min_k

label: Minimum k-mer length at right end of reads used for trimming
type: basic:integer
disabled: preprocessing.adapters.length === 0 && preprocessing.custom_adapter_sequences.length === 0
default: 11

preprocessing.hamming_distance

label: Maximum Hamming distance for k-mers
type: basic:integer
default: 1

preprocessing.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
default: -1

preprocessing.trim_quality

label: Quality below which to trim reads from the right end
type: basic:integer
description: Phred algorithm is used, which is more accurate than naive trimming.
default: 10

preprocessing.min_length

label: Minimum read length
type: basic:integer
description: Reads shorter than minimum read length after trimming are discarded.
default: 20

quantification.seq_bias

label: Perform sequence-specific bias correction
type: basic:boolean
default: True

quantification.gc_bias

label: Perform fragment GC bias correction.
type: basic:boolean
default: True

quantification.consensus_slack

label: Consensus slack
type: basic:decimal
description: The amount of slack allowed in the quasi-mapping consensus mechanism. Normally, a transcript must cover all hits to be considered for mapping. If this is set to a fraction, X, greater than 0 (and in [0,1)), then a transcript can fail to cover up to (100 * X)% of the hits before it is discounted as a mapping candidate. The default value of this option is 0.2 in selective alignment mode and 0 otherwise.
required: False

quantification.min_score_fraction

label: Minimum alignment score fraction
type: basic:decimal
description: The fraction of the optimal possible alignment score that a mapping must achieve in order to be considered valid - should be in (0,1].
default: 0.65

quantification.range_factorization_bins

label: Range factorization bins
type: basic:integer
description: Factorizes the likelihood used in quantification by adopting a new notion of equivalence classes based on the conditional probabilities with which fragments are generated from different transcripts. This is a more fine-grained factorization than the normal rich equivalence classes. The default value (4) corresponds to the default used in Zakeri et al. 2017 and larger values imply a more fine-grained factorization. If range factorization is enabled, a common value to select for this parameter is 4. A value of 0 signifies the use of basic rich equivalence classes.
default: 4

quantification.min_assigned_frag

label: Minimum number of assigned fragments
type: basic:integer
description: The minimum number of fragments that must be assigned to the transcriptome for quantification to proceed.
default: 10

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 10000000

downsampling.seed

label: Seed
type: basic:integer
default: 11

downsampling.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads [0 - 1.0] from the original input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

BBDuk - Salmon - QC (single-end)¶

data:workflow:rnaseq:salmonworkflow-bbduk-salmon-qc-single (data:reads:fastq:single reads, data:index:salmon salmon_index, data:index:star genome, data:annotation annotation, data:index:star rrna_reference, data:index:star globin_reference, basic:boolean show_advanced, list:data:seq:nucleotide adapters, list:basic:string custom_adapter_sequences, basic:integer kmer_length, basic:integer min_k, basic:integer hamming_distance, basic:integer maxns, basic:integer trim_quality, basic:integer min_length, basic:boolean seq_bias, basic:boolean gc_bias, basic:decimal consensus_slack, basic:decimal min_score_fraction, basic:integer range_factorization_bins, basic:integer min_assigned_frag, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass)[Source: v3.0.1]

Alignment-free RNA-seq pipeline. Salmon tool and tximport package are used in quantification step to produce gene-level abundance estimates. rRNA and globin-sequence contamination rate in the sample is determined using STAR aligner. Quality-trimmed reads are down-sampled (using Seqtk tool) and aligned to the genome, rRNA and globin reference sequences. The rRNA and globin-sequence alignment rates indicate the percentage of the reads in the sample that are of rRNA and globin origin, respectively. Alignment of down-sampled data to a whole genome reference sequence is used to produce an alignment file suitable for Samtools and QoRTs QC analysis. Per-sample analysis results and QC data is summarized by the MultiQC tool.

reads

label: Select sample(s)
type: data:reads:fastq:single

salmon_index

label: Salmon index
type: data:index:salmon

genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation

rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

preprocessing.adapters

label: Adapters
type: list:data:seq:nucleotide
required: False

preprocessing.custom_adapter_sequences

label: Custom adapter sequences [literal]
type: list:basic:string
description: Custom adapter sequences can be specified by inputting them one by one and pressing Enter after each sequence.
required: False
default: []

preprocessing.kmer_length

label: K-mer length
type: basic:integer
description: K-mer length must be smaller or equal to the length of adapters.
default: 23

preprocessing.min_k

label: Minimum k-mer length at right end of reads used for trimming
type: basic:integer
disabled: preprocessing.adapters.length === 0 && preprocessing.custom_adapter_sequences.length === 0
default: 11

preprocessing.hamming_distance

label: Maximum Hamming distance for k-mers
type: basic:integer
default: 1

preprocessing.maxns

label: Max Ns after trimming [maxns=-1]
type: basic:integer
description: If non-negative, reads with more Ns than this (after trimming) will be discarded.
default: -1

preprocessing.trim_quality

label: Quality below which to trim reads from the right end
type: basic:integer
description: Phred algorithm is used, which is more accurate than naive trimming.
default: 10

preprocessing.min_length

label: Minimum read length
type: basic:integer
description: Reads shorter than minimum read length after trimming are discarded.
default: 20

quantification.seq_bias

label: Perform sequence-specific bias correction
type: basic:boolean
default: True

quantification.gc_bias

label: Perform fragment GC bias correction.
type: basic:boolean
default: False

quantification.consensus_slack

label: Consensus slack
type: basic:decimal
description: The amount of slack allowed in the quasi-mapping consensus mechanism. Normally, a transcript must cover all hits to be considered for mapping. If this is set to a fraction, X, greater than 0 (and in [0,1)), then a transcript can fail to cover up to (100 * X)% of the hits before it is discounted as a mapping candidate. The default value of this option is 0.2 in selective alignment mode and 0 otherwise.
required: False

quantification.min_score_fraction

label: Minimum alignment score fraction
type: basic:decimal
description: The fraction of the optimal possible alignment score that a mapping must achieve in order to be considered valid - should be in (0,1].
default: 0.65

quantification.range_factorization_bins

label: Range factorization bins
type: basic:integer
description: Factorizes the likelihood used in quantification by adopting a new notion of equivalence classes based on the conditional probabilities with which fragments are generated from different transcripts. This is a more fine-grained factorization than the normal rich equivalence classes. The default value (4) corresponds to the default used in Zakeri et al. 2017 and larger values imply a more fine-grained factorization. If range factorization is enabled, a common value to select for this parameter is 4. A value of 0 signifies the use of basic rich equivalence classes.
default: 4

quantification.min_assigned_frag

label: Minimum number of assigned fragments
type: basic:integer
description: The minimum number of fragments that must be assigned to the transcriptome for quantification to proceed.
default: 10

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 10000000

downsampling.seed

label: Seed
type: basic:integer
default: 11

downsampling.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads [0 - 1.0] from the original input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

BED file¶

data:bedupload-bed (basic:file src, basic:string species, basic:string build)[Source: v1.4.1]

Import a BED file (.bed) which is a tab-delimited text file that defines a feature track. It can have any file extension, but .bed is recommended. The BED file format is described on the [UCSC Genome Bioinformatics web site](http://genome.ucsc.edu/FAQ/FAQformat#format1).

src

label: BED file
type: basic:file
description: Upload BED file annotation track. The first three required BED fields are chrom, chromStart and chromEnd.
required: True
validate_regex: \.(bed|narrowPeak)$

species

label

Species

type

basic:string

description

Species latin name.

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Genome build
type: basic:string

bed

label: BED file
type: basic:file

bed_jbrowse

label: Bgzip bed file for JBrowse
type: basic:file

tbi_jbrowse

label: Bed file index for Jbrowse
type: basic:file

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BEDPE file¶

data:bedpe:upload-bedpe (basic:file src, basic:string species, basic:string build)[Source: v1.2.1]

Upload BEDPE files.

src

label: Select BEDPE file to upload
type: basic:file
required: True
hidden: False

species

label

Species

type

basic:string

required

True

hidden

False

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Build
type: basic:string
required: True
hidden: False

bedpe

label: BEDPE file
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

BWA ALN¶

data:alignment:bam:bwaalnalignment-bwa-aln (data:index:bwa genome, data:reads:fastq reads, basic:integer q, basic:boolean use_edit, basic:integer edit_value, basic:decimal fraction, basic:boolean seeds, basic:integer seed_length, basic:integer seed_dist)[Source: v2.3.1]

Read aligner for mapping low-divergent sequences against a large reference genome. Designed for Illumina sequence reads up to 100bp.

genome

label: Reference genome
type: data:index:bwa

reads

label: Reads
type: data:reads:fastq

q

label: Quality threshold
type: basic:integer
description: Parameter for dynamic read trimming.
default: 0

use_edit

label: Use maximum edit distance (excludes fraction of missing alignments)
type: basic:boolean
default: False

edit_value

label: Maximum edit distance
type: basic:integer
hidden: !use_edit
default: 5

fraction

label: Fraction of missing alignments
type: basic:decimal
description: The fraction of missing alignments given 2% uniform base error rate. The maximum edit distance is automatically chosen for different read lengths.
hidden: use_edit
default: 0.04

seeds

label: Use seeds
type: basic:boolean
default: False

seed_length

label: Seed length
type: basic:integer
description: Take the first X subsequence as seed. If X is larger than the query sequence, seeding will be disabled. For long reads, this option is typically ranged from 25 to 35 for value 2 in seed maximum edit distance.
hidden: !seeds
default: 35

seed_dist

label: Seed maximum edit distance
type: basic:integer
hidden: !seeds
default: 2

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

unmapped

label: Unmapped reads
type: basic:file
required: False

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BWA MEM¶

data:alignment:bam:bwamemalignment-bwa-mem (data:index:bwa genome, data:reads:fastq reads, basic:integer seed_l, basic:integer band_w, basic:decimal re_seeding, basic:boolean m, basic:integer match, basic:integer missmatch, basic:integer gap_o, basic:integer gap_e, basic:integer clipping, basic:integer unpaired_p, basic:boolean report_all, basic:integer report_tr)[Source: v3.3.2]

BWA MEM is a read aligner for mapping low-divergent sequences against a large reference genome. Designed for longer sequences ranged from 70bp to 1Mbp. The algorithm works by seeding alignments with maximal exact matches (MEMs) and then extending seeds with the affine-gap Smith-Waterman algorithm (SW). See [here](http://bio-bwa.sourceforge.net/) for more information.

genome

label: Reference genome
type: data:index:bwa

reads

label: Reads
type: data:reads:fastq

seed_l

label: Minimum seed length
type: basic:integer
description: Minimum seed length. Matches shorter than minimum seed length will be missed. The alignment speed is usually insensitive to this value unless it significantly deviates from 20.
default: 19

band_w

label: Band width
type: basic:integer
description: Gaps longer than this will not be found.
default: 100

re_seeding

label: Re-seeding factor
type: basic:decimal
description: Trigger re-seeding for a MEM longer than minSeedLen*FACTOR. This is a key heuristic parameter for tuning the performance. Larger value yields fewer seeds, which leads to faster alignment speed but lower accuracy.
default: 1.5

m

label: Mark shorter split hits as secondary
type: basic:boolean
description: Mark shorter split hits as secondary (for Picard compatibility)
default: False

scoring.match

label: Score of a match
type: basic:integer
default: 1

scoring.missmatch

label: Mismatch penalty
type: basic:integer
default: 4

scoring.gap_o

label: Gap open penalty
type: basic:integer
default: 6

scoring.gap_e

label: Gap extension penalty
type: basic:integer
default: 1

scoring.clipping

label: Clipping penalty
type: basic:integer
description: Clipping is applied if final alignment score is smaller than (best score reaching the end of query) - (Clipping penalty)
default: 5

scoring.unpaired_p

label: Penalty for an unpaired read pair
type: basic:integer
description: Affinity to force pair. Score: scoreRead1+scoreRead2-Penalty
default: 9

reporting.report_all

label: Report all found alignments
type: basic:boolean
description: Output all found alignments for single-end or unpaired paired-end reads. These alignments will be flagged as secondary alignments.
default: False

reporting.report_tr

label: Report threshold score
type: basic:integer
description: Don’t output alignment with score lower than defined number. This option only affects output.
default: 30

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

unmapped

label: Unmapped reads
type: basic:file
required: False

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BWA SW¶

data:alignment:bam:bwaswalignment-bwa-sw (data:index:bwa genome, data:reads:fastq reads, basic:integer match, basic:integer missmatch, basic:integer gap_o, basic:integer gap_e)[Source: v2.3.1]

Read aligner for mapping low-divergent sequences against a large reference genome. Designed for longer sequences ranged from 70bp to 1Mbp. The paired-end mode only works for reads Illumina short-insert libraries.

genome

label: Reference genome
type: data:index:bwa

reads

label: Reads
type: data:reads:fastq

match

label: Score of a match
type: basic:integer
default: 1

missmatch

label: Mismatch penalty
type: basic:integer
default: 3

gap_o

label: Gap open penalty
type: basic:integer
default: 5

gap_e

label: Gap extension penalty
type: basic:integer
default: 2

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

unmapped

label: Unmapped reads
type: basic:file
required: False

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

BWA genome index¶

data:index:bwa:bwa-index (data:seq:nucleotide ref_seq)[Source: v1.1.1]

Create BWA genome index.

ref_seq

label: Reference sequence (nucleotide FASTA)
type: data:seq:nucleotide
required: True
hidden: False

index

label: BWA index
type: basic:dir
required: True
hidden: False

fastagz

label: FASTA file (compressed)
type: basic:file
required: True
hidden: False

fasta

label: FASTA file
type: basic:file
required: True
hidden: False

fai

label: FASTA file index
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Bam split¶

data:alignment:bam:primarybam-split (data:alignment:bam bam, data:sam:header header, data:sam:header header2)[Source: v0.6.1]

Split hybrid bam file into two bam files.

bam

label: Hybrid alignment bam
type: data:alignment:bam

header

label: Primary header sam file (optional)
type: data:sam:header
description: If no header file is provided, the headers will be extracted from the hybrid alignment bam file.
required: False

header2

label: Secondary header sam file (optional)
type: data:sam:header
description: If no header file is provided, the headers will be extracted from the hybrid alignment bam file.
required: False

bam

label: Uploaded file
type: basic:file

bai

label: Index BAI
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Bamclipper¶

data:alignment:bam:bamclipped:bamclipper (data:alignment:bam alignment, data:bedpe bedpe, basic:boolean skip)[Source: v1.2.1]

Remove primer sequence from BAM alignments by soft-clipping. This process is a wrapper for bamclipper which can be found at https://github.com/tommyau/bamclipper.

alignment

label: Alignment BAM file
type: data:alignment:bam
required: True
hidden: False

bedpe

label: BEDPE file
type: data:bedpe
required: False
hidden: False

skip

label: Skip Bamclipper step
type: basic:boolean
description: Use this option to skip Bamclipper step.
required: True
hidden: False
default: False

bam

label: Clipped BAM file
type: basic:file
required: True
hidden: False

bai

label: Index of clipped BAM file
type: basic:file
required: True
hidden: False

stats

label: Alignment statistics
type: basic:file
required: True
hidden: False

bigwig

label: BigWig file
type: basic:file
required: False
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Bamliquidator¶

data:bam:plot:bamliquidatorbamliquidator (basic:string analysis_type, list:data:alignment:bam bam, basic:string cell_type, basic:integer bin_size, data:annotation:gtf regions_gtf, data:bed regions_bed, basic:integer extension, basic:string sense, basic:boolean skip_plot, list:basic:string black_list, basic:integer threads)[Source: v0.3.1]

Set of tools for analyzing the density of short DNA sequence read alignments in the BAM file format.

analysis_type

label

Analysis type

type

basic:string

default

bin

choices

Bin mode: bin
Region mode: region
BED mode: bed

bam

label: BAM File
type: list:data:alignment:bam

cell_type

label: Cell type
type: basic:string
default: cell_type

bin_size

label: Bin size
type: basic:integer
description: Number of base pairs in each bin. The smaller the bin size the longer the runtime and the larger the data files. Default is 100000.
required: False
hidden: analysis_type != ‘bin’

regions_gtf

label: Region gff file / Annotation file (.gff|.gtf)
type: data:annotation:gtf
required: False
hidden: analysis_type != ‘region’

regions_bed

label: Region bed file / Annotation file (.bed)
type: data:bed
required: False
hidden: analysis_type != ‘bed’

extension

label: Extension
type: basic:integer
description: Extends reads by number of bp
default: 200

sense

label

Mapping strand to gff file

type

basic:string

default

.

choices

Forward: +
Reverse: -
Both: .

skip_plot

label: Skip plot
type: basic:boolean
required: False

black_list

label: Black list
type: list:basic:string
description: One or more chromosome patterns to skip during bin liquidation. Default is to skip any chromosomes that contain any of the following substrings chrUn _random Zv9_ _hap.
required: False

threads

label: Threads
type: basic:integer
description: Number of threads to run concurrently during liquidation.
default: 1

analysis_type

label: Analysis type
type: basic:string
hidden: True

output_dir

label: Output directory
type: basic:file

counts

label: Counts HDF5 file
type: basic:file

matrix

label: Matrix file
type: basic:file
required: False
hidden: analysis_type != ‘region’

summary

label: Summary file
type: basic:file:html
required: False
hidden: analysis_type != ‘bin’

Bamplot¶

data:bam:plot:bamplotbamplot (basic:string genome, data:annotation:gtf input_gff, basic:string input_region, list:data:alignment:bam bam, basic:integer stretch_input, basic:string color, basic:string sense, basic:integer extension, basic:boolean rpm, basic:string yscale, list:basic:string names, basic:string plot, basic:string title, basic:string scale, list:data:bed bed, basic:boolean multi_page)[Source: v1.4.1]

Plot a single locus from a bam.

genome

label

Genome

type

basic:string

choices

HG19: HG19
HG18: HG18
MM8: MM8
MM9: MM9
MM10: MM10
RN6: RN6
RN4: RN4

input_gff

label: Region string
type: data:annotation:gtf
description: Enter .gff file.
required: False

input_region

label: Region string
type: basic:string
description: Enter genomic region e.g. chr1:+:1-1000.
required: False

bam

label: Bam
type: list:data:alignment:bam
description: bam to plot from
required: False

stretch_input

label: Stretch-input
type: basic:integer
description: Stretch the input regions to a minimum length in bp, e.g. 10000 (for 10kb).
required: False

color

label: Color
type: basic:string
description: Enter a colon separated list of colors e.g. 255,0,0:255,125,0, default samples the rainbow.
default: 255,0,0:255,125,0

sense

label

Sense

type

basic:string

description

Map to forward, reverse or’both strands. Default maps to both.

default

both

choices

Forward: forward
Reverse: reverse
Both: both

extension

label: Extension
type: basic:integer
description: Extends reads by n bp. Default value is 200bp.
default: 200

rpm

label: rpm
type: basic:boolean
description: Normalizes density to reads per million (rpm) Default is False.
required: False

yscale

label

y scale

type

basic:string

description

Choose either relative or uniform y axis scaling. Default is relative scaling.

default

relative

choices

relative: relative
uniform: uniform

names

label: Names
type: list:basic:string
description: Enter a comma separated list of names for your bams.
required: False

plot

label

Single or multiple polt

type

basic:string

description

Choose either all lines on a single plot or multiple plots.

default

merge

choices

single: single
multiple: multiple
merge: merge

title

label: Title
type: basic:string
description: Specify a title for the output plot(s), default will be the coordinate region.
default: output

scale

label: Scale
type: basic:string
description: Enter a comma separated list of multiplicative scaling factors for your bams. Default is none.
required: False

bed

label: Bed
type: list:data:bed
description: Add a space-delimited list of bed files to plot.
required: False

multi_page

label: Multi page
type: basic:boolean
description: If flagged will create a new pdf for each region.
default: False

plot

label: region plot
type: basic:file

BaseQualityScoreRecalibrator¶

data:alignment:bam:bqsr:bqsr (data:alignment:bam bam, data:seq:nucleotide reference, list:data:variants:vcf known_sites, data:bed intervals, basic:string read_group, basic:string validation_stringency)[Source: v2.1.1]

A two pass process of BaseRecalibrator and ApplyBQSR from GATK. See GATK website for more information on BaseRecalibrator. It is possible to modify read group using GATK’s AddOrReplaceGroups through Replace read groups in BAM (``read_group``) input field.

bam

label: BAM file containing reads
type: data:alignment:bam
required: True
hidden: False

reference

label: Reference genome file
type: data:seq:nucleotide
required: True
hidden: False

known_sites

label: List of known sites of variation
type: list:data:variants:vcf
required: True
hidden: False

intervals

label: One or more genomic intervals over which to operate.
type: data:bed
description: This field is optional, but it can speed up the process by restricting calculations to specific genome regions.
required: False
hidden: False

read_group

label: Replace read groups in BAM
type: basic:string
description: Replace read groups in a BAM file.This argument enables the user to replace all read groups in the INPUT file with a single new read group and assign all reads to this read group in the OUTPUT BAM file. Addition or replacement is performed using Picard’s AddOrReplaceReadGroups tool. Input should take the form of -name=value delimited by a “;”, e.g. “-ID=1;-LB=GENIALIS;-PL=ILLUMINA;-PU=BARCODE;-SM=SAMPLENAME1”. See tool’s documentation for more information on tag names. Note that PL, LB, PU and SM are require fields. See caveats of rewriting read groups in the documentation.
required: True
hidden: False
default

validation_stringency

label

Validation stringency

type

basic:string

description

Validation stringency for all SAM files read by this program. Setting stringency to SILENT can improve performance when processing a BAM file in which variable-length data (read, qualities, tags) do not otherwise need to be decoded. Default is STRICT. This setting is used in BaseRecalibrator and ApplyBQSR processes.

required

True

hidden

False

default

STRICT

choices

STRICT: STRICT
LENIENT: LENIENT
SILENT: SILENT

bam

label: Base quality score recalibrated BAM file
type: basic:file
required: True
hidden: False

bai

label: Index of base quality score recalibrated BAM file
type: basic:file
required: True
hidden: False

stats

label: Alignment statistics
type: basic:file
required: True
hidden: False

bigwig

label: BigWig file
type: basic:file
required: False
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

recal_table

label: Recalibration tabled
type: basic:file
required: True
hidden: False

BaseSpace file¶

data:filebasespace-file-import (basic:string file_id, basic:secret access_token_secret)[Source: v1.2.1]

Import a file from Illumina BaseSpace.

file_id

label: BaseSpace file ID
type: basic:string

access_token_secret

label: BaseSpace access token
type: basic:secret
description: BaseSpace access token secret handle needed to download the file.

file

label: File
type: basic:file

Bedtools bamtobed¶

data:bedpe:bedtools-bamtobed (data:alignment:bam alignment)[Source: v1.1.1]

Takes in a BAM file and calculates a normalization factor in BEDPE format. Done by sorting with Samtools and transformed with Bedtools.

alignment

label: Alignment BAM file
type: data:alignment:bam
required: True
hidden: False

bedpe

label: BEDPE file
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Bisulfite conversion rate¶

data:wgbs:bsrate:bs-conversion-rate (data:alignment:bam:walt mr, basic:boolean skip, data:seq:nucleotide sequence, basic:boolean count_all, basic:integer read_length, basic:decimal max_mismatch, basic:boolean a_rich)[Source: v1.1.1]

Estimate bisulfite conversion rate in a control set. The program bsrate included in [Methpipe] (https://github.com/smithlabcode/methpipe) will estimate the bisulfite conversion rate.

mr

label: Aligned reads from bisulfite sequencing
type: data:alignment:bam:walt
description: Bisulfite specifc alignment such as WALT is required as .mr file type is used. Duplicatesshould be removed to reduce any bias introduced by incomplete conversion on PCR duplicatereads.
required: True
hidden: False

skip

label: Skip Bisulfite conversion rate step
type: basic:boolean
description: Bisulfite conversion rate step can be skipped.
required: True
hidden: False
default: False

sequence

label: Unmethylated control sequence
type: data:seq:nucleotide
description: Separate unmethylated control sequence FASTA file is required to estimate bisulfiteconversion rate.
required: False
hidden: False

count_all

label: Count all cytosines including CpGs
type: basic:boolean
required: True
hidden: False
default: True

read_length

label: Average read length
type: basic:integer
required: True
hidden: False
default: 150

max_mismatch

label: Maximum fraction of mismatches
type: basic:decimal
required: False
hidden: False

a_rich

label: Reads are A-rich
type: basic:boolean
required: True
hidden: False
default: False

report

label: Bisulfite conversion rate report
type: basic:file
required: True
hidden: False

Bowtie (Dicty)¶

data:alignment:bam:bowtie1alignment-bowtie (data:index:bowtie genome, data:reads:fastq reads, basic:string mode, basic:integer m, basic:integer l, basic:boolean use_se, basic:integer trim_5, basic:integer trim_3, basic:integer trim_nucl, basic:integer trim_iter, basic:string r)[Source: v2.3.1]

An ultrafast memory-efficient short read aligner.

genome

label: Reference genome
type: data:index:bowtie

reads

label: Reads
type: data:reads:fastq

mode

label

Alignment mode

type

basic:string

description

When the -n option is specified (which is the default), bowtie determines which alignments are valid according to the following policy, which is similar to Maq’s default policy. 1. Alignments may have no more than N mismatches (where N is a number 0-3, set with -n) in the first L bases (where L is a number 5 or greater, set with -l) on the high-quality (left) end of the read. The first L bases are called the “seed”. 2. The sum of the Phred quality values at all mismatched positions (not just in the seed) may not exceed E (set with -e). Where qualities are unavailable (e.g. if the reads are from a FASTA file), the Phred quality defaults to 40. In -v mode, alignments may have no more than V mismatches, where V may be a number from 0 through 3 set using the -v option. Quality values are ignored. The -v option is mutually exclusive with the -n option.

default

-n

choices

Use qualities (-n): -n
Use mismatches (-v): -v

m

label: Allowed mismatches
type: basic:integer
description: When used with “Use qualities (-n)” it is the maximum number of mismatches permitted in the “seed”, i.e. the first L base pairs of the read (where L is set with -l/–seedlen). This may be 0, 1, 2 or 3 and the default is 2 When used with “Use mismatches (-v)” report alignments with at most <int> mismatches.
default: 2

l

label: Seed length (for -n only)
type: basic:integer
description: Only for “Use qualities (-n)”. Seed length (-l) is the number of bases on the high-quality end of the read to which the -n ceiling applies. The lowest permitted setting is 5 and the default is 28. bowtie is faster for larger values of -l.
default: 28

use_se

label: Map as single-ended (for paired end reads only)
type: basic:boolean
description: If this option is selected paired-end reads will be mapped as single-ended.
default: False

start_trimming.trim_5

label: Bases to trim from 5’
type: basic:integer
description: Number of bases to trim from from 5’ (left) end of each read before alignment
default: 0

start_trimming.trim_3

label: Bases to trim from 3’
type: basic:integer
description: Number of bases to trim from from 3’ (right) end of each read before alignment
default: 0

trimming.trim_nucl

label: Bases to trim
type: basic:integer
description: Number of bases to trim from 3’ end in each iteration.
default: 2

trimming.trim_iter

label: Iterations
type: basic:integer
description: Number of iterations.
default: 0

reporting.r

label

Reporting mode

type

basic:string

description

Report up to <int> valid alignments per read or pair (-k) (default: 1). Validity of alignments is determined by the alignment policy (combined effects of -n, -v, -l, and -e). If more than one valid alignment exists and the –best and –strata options are specified, then only those alignments belonging to the best alignment “stratum” will be reported. Bowtie is designed to be very fast for small -k but bowtie can become significantly slower as -k increases. If you would like to use Bowtie for larger values of -k, consider building an index with a denser suffix-array sample, i.e. specify a smaller -o/–offrate when invoking bowtie-build for the relevant index (see the Performance tuning section for details).

default

-a -m 1 --best --strata

choices

Report unique alignments: -a -m 1 --best --strata
Report all alignments: -a --best
Report all alignments in the best stratum: -a --best --strata

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

unmapped

label: Unmapped reads
type: basic:file
required: False

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Bowtie genome index¶

data:index:bowtie:bowtie-index (data:seq:nucleotide ref_seq)[Source: v1.1.1]

Create Bowtie genome index.

ref_seq

label: Reference sequence (nucleotide FASTA)
type: data:seq:nucleotide
required: True
hidden: False

index

label: Bowtie index
type: basic:dir
required: True
hidden: False

fastagz

label: FASTA file (compressed)
type: basic:file
required: True
hidden: False

fasta

label: FASTA file
type: basic:file
required: True
hidden: False

fai

label: FASTA file index
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Bowtie2¶

data:alignment:bam:bowtie2alignment-bowtie2 (data:index:bowtie2 genome, data:reads:fastq reads, basic:string mode, basic:string speed, basic:boolean use_se, basic:boolean discordantly, basic:boolean rep_se, basic:integer minins, basic:integer maxins, basic:boolean no_overlap, basic:boolean dovetail, basic:integer N, basic:integer L, basic:integer gbar, basic:string mp, basic:string rdg, basic:string rfg, basic:string score_min, basic:integer trim_5, basic:integer trim_3, basic:integer trim_iter, basic:integer trim_nucl, basic:string rep_mode, basic:integer k_reports, basic:boolean no_unal, basic:integer bw_binsize, basic:integer bw_timeout)[Source: v2.5.1]

Bowtie is an ultrafast, memory-efficient short read aligner. It aligns short DNA sequences (reads) to the human genome at a rate of over 25 million 35-bp reads per hour. Bowtie indexes the genome with a Burrows-Wheeler index to keep its memory footprint small–typically about 2.2 GB for the human genome (2.9 GB for paired-end). See [here](http://bowtie-bio.sourceforge.net/index.shtml) for more information.

genome

label: Reference genome
type: data:index:bowtie2

reads

label: Reads
type: data:reads:fastq

mode

label

Alignment mode

type

basic:string

description

End to end: Bowtie 2 requires that the entire read align from one end to the other, without any trimming (or “soft clipping”) of characters from either end. local: Bowtie 2 does not require that the entire read align from one end to the other. Rather, some characters may be omitted (“soft clipped”) from the ends in order to achieve the greatest possible alignment score.

default

--end-to-end

choices

end to end mode: --end-to-end
local: --local

speed

label

Speed vs. Sensitivity

type

basic:string

description

A quick setting for aligning fast or accurately. This option is a shortcut for parameters as follows: For –end-to-end: –very-fast -D 5 -R 1 -N 0 -L 22 -i S,0,2.50 –fast -D 10 -R 2 -N 0 -L 22 -i S,0,2.50 –sensitive -D 15 -R 2 -N 0 -L 22 -i S,1,1.15 (default) –very-sensitive -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 For –local: –very-fast-local -D 5 -R 1 -N 0 -L 25 -i S,1,2.00 –fast-local -D 10 -R 2 -N 0 -L 22 -i S,1,1.75 –sensitive-local -D 15 -R 2 -N 0 -L 20 -i S,1,0.75 (default) –very-sensitive-local -D 20 -R 3 -N 0 -L 20 -i S,1,0.50

required

False

choices

Very fast: --very-fast
Fast: --fast
Sensitive: --sensitive
Very sensitive: --very-sensitive

PE_options.use_se

label: Map as single-ended (for paired-end reads only)
type: basic:boolean
description: If this option is selected paired-end reads will be mapped as single-ended and other paired-end options are ignored.
default: False

PE_options.discordantly

label: Report discordantly matched read
type: basic:boolean
description: If both mates have unique alignments, but the alignments do not match paired-end expectations (orientation and relative distance) then alignment will be reported. Useful for detecting structural variations.
default: True

PE_options.rep_se

label: Report single ended
type: basic:boolean
description: If paired alignment can not be found Bowtie2 tries to find alignments for the individual mates.
default: True

PE_options.minins

label: Minimal distance
type: basic:integer
description: The minimum fragment length for valid paired-end alignments. 0 imposes no minimum.
default: 0

PE_options.maxins

label: Maximal distance
type: basic:integer
description: The maximum fragment length for valid paired-end alignments.
default: 500

PE_options.no_overlap

label: Not concordant when mates overlap
type: basic:boolean
description: When true, it is considered not concordant when mates overlap at all. Defaul is false.
default: False

PE_options.dovetail

label: Dovetail
type: basic:boolean
description: If the mates “dovetail”, that is if one mate alignment extends past the beginning of the other such that the wrong mate begins upstream, consider that to be concordant. Default: mates cannot dovetail in a concordant alignment.
default: False

alignment_options.N

label: Number of mismatches allowed in seed alignment (N)
type: basic:integer
description: Sets the number of mismatches to allowed in a seed alignment during multiseed alignment. Can be set to 0 or 1. Setting this higher makes alignment slower (often much slower) but increases sensitivity. Default: 0.
required: False

alignment_options.L

label: Length of seed substrings (L)
type: basic:integer
description: Sets the length of the seed substrings to align during multiseed alignment. Smaller values make alignment slower but more sensitive. Default: the –sensitive preset is used by default for end-to-end alignment and –sensitive-local for local alignment. See documentation for details.
required: False

alignment_options.gbar

label: Disallow gaps within positions (gbar)
type: basic:integer
description: Disallow gaps within <int> positions of the beginning or end of the read. Default: 4.
required: False

alignment_options.mp

label: Maximal and minimal mismatch penalty (mp)
type: basic:string
description: Sets the maximum (MX) and minimum (MN) mismatch penalties, both integers. A number less than or equal to MX and greater than or equal to MN is subtracted from the alignment score for each position where a read character aligns to a reference character, the characters do not match, and neither is an N. If –ignore-quals is specified, the number subtracted quals MX. Otherwise, the number subtracted is MN + floor((MX-MN)(MIN(Q, 40.0)/40.0)) where Q is the Phred quality value. Default for MX, MN: 6,2.
required: False

alignment_options.rdg

label: Set read gap open and extend penalties (rdg)
type: basic:string
description: Sets the read gap open (<int1>) and extend (<int2>) penalties. A read gap of length N gets a penalty of <int1> + N * <int2>. Default: 5,3.
required: False

alignment_options.rfg

label: Set reference gap open and close penalties (rfg)
type: basic:string
description: Sets the reference gap open (<int1>) and extend (<int2>) penalties. A reference gap of length N gets a penalty of <int1> + N * <int2>. Default: 5,3.
required: False

alignment_options.score_min

label: Minimum alignment score needed for “valid” alignment (score_min)
type: basic:string
description: Sets a function governing the minimum alignment score needed for an alignment to be considered “valid” (i.e. good enough to report). This is a function of read length. For instance, specifying L,0,-0.6 sets the minimum-score function to f(x) = 0 + -0.6 * x, where x is the read length. The default in –end-to-end mode is L,-0.6,-0.6 and the default in –local mode is G,20,8.
required: False

start_trimming.trim_5

label: Bases to trim from 5’
type: basic:integer
description: Number of bases to trim from from 5’ (left) end of each read before alignment
default: 0

start_trimming.trim_3

label: Bases to trim from 3’
type: basic:integer
description: Number of bases to trim from from 3’ (right) end of each read before alignment
default: 0

trimming.trim_iter

label: Iterations
type: basic:integer
description: Number of iterations.
default: 0

trimming.trim_nucl

label: Bases to trim
type: basic:integer
description: Number of bases to trim from 3’ end in each iteration.
default: 2

reporting.rep_mode

label

Report mode

type

basic:string

description

Default mode: search for multiple alignments, report the best one; -k mode: search for one or more alignments, report each; -a mode: search for and report all alignments

default

def

choices

Default mode: def
-k mode: k
-a mode (very slow): a

reporting.k_reports

label: Number of reports (for -k mode only)
type: basic:integer
description: Searches for at most X distinct, valid alignments for each read. The search terminates when it can’t find more distinct valid alignments, or when it finds X, whichever happens first. default: 5
default: 5

output_opts.no_unal

label: Suppress SAM records for unaligned reads
type: basic:boolean
description: When true, suppress SAM records for unaligned reads. Default is false.
default: False

misc_opts.bw_binsize

label: BigWig bin size
type: basic:integer
description: Size of the bins, in bases, for the output of the bigwig/bedgraph file. Default is 50.
default: 50

misc_opts.bw_timeout

label: BigWig timeout (s)
type: basic:integer
description: Time, in seconds, before creation of BigWig file is stopped. Default is 480 seconds.
default: 480

bam

label: Alignment file
type: basic:file
description: Position sorted alignment

bai

label: Index BAI
type: basic:file

unmapped

label: Unmapped reads
type: basic:file
required: False

stats

label: Statistics
type: basic:file

bigwig

label: BigWig file
type: basic:file
required: False

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Bowtie2 genome index¶

data:index:bowtie2:bowtie2-index (data:seq:nucleotide ref_seq)[Source: v1.1.1]

Create Bowtie2 genome index.

ref_seq

label: Reference sequence (nucleotide FASTA)
type: data:seq:nucleotide
required: True
hidden: False

index

label: Bowtie2 index
type: basic:dir
required: True
hidden: False

fastagz

label: FASTA file (compressed)
type: basic:file
required: True
hidden: False

fasta

label: FASTA file
type: basic:file
required: True
hidden: False

fai

label: FASTA file index
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Cell Ranger Count¶

data:scexpression:10x:cellranger-count (data:screads:10x: reads, data:genomeindex:10x: genome_index, basic:string chemistry, basic:integer trim_r1, basic:integer trim_r2, basic:integer expected_cells, basic:integer force_cells)[Source: v1.1.1]

Perform gene expression analysis. Generate single cell feature counts for a single library. https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/using/count

reads

label: 10x reads data object
type: data:screads:10x:
required: True
hidden: False

genome_index

label: 10x genome index data object
type: data:genomeindex:10x:
required: True
hidden: False

chemistry

label

Chemistry

type

basic:string

description

Assay configuration. By default the assay configuration is detected automatically, which is the recommended mode. You should only specify chemistry if there is an error in automatic detection.

required

False

hidden

False

default

auto

choices

auto: auto
threeprime: Single Cell 3'
fiveprime: Single Cell 5'
SC3Pv1: Single Cell 3' v1
SC3Pv2: Single Cell 3' v2
SC3Pv3: Single Cell 3' v3
C5P-PE: Single Cell 5' paired-end
SC5P-R2: Single Cell 5' R2-only

trim_r1

label: Trim R1
type: basic:integer
description: Hard-trim the input R1 sequence to this length. Note that the length includes the Barcode and UMI sequences so do not set this below 26 for Single Cell 3’ v2 or Single Cell 5’. This and “Trim R2” are useful for determining the optimal read length for sequencing.
required: False
hidden: False

trim_r2

label: Trim R2
type: basic:integer
description: Hard-trim the input R2 sequence to this length.
required: False
hidden: False

expected_cells

label: Expected number of recovered cells
type: basic:integer
required: True
hidden: False
default: 3000

force_cells

label: Force cell number
type: basic:integer
description: Force pipeline to use this number of cells, bypassing the cell detection algorithm. Use this if the number of cells estimated by Cell Ranger is not consistent with the barcode rank plot.
required: False
hidden: False

matrix_filtered

label: Matrix (filtered)
type: basic:file
required: True
hidden: False

genes_filtered

label: Genes (filtered)
type: basic:file
required: True
hidden: False

barcodes_filtered

label: Barcodes (filtered)
type: basic:file
required: True
hidden: False

matrix_raw

label: Matrix (raw)
type: basic:file
required: True
hidden: False

genes_raw

label: Genes (raw)
type: basic:file
required: True
hidden: False

barcodes_raw

label: Barcodes (raw)
type: basic:file
required: True
hidden: False

report

label: Report
type: basic:file:html
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

source

label: Gene ID source
type: basic:string
required: True
hidden: False

Cell Ranger Mkref¶

data:genomeindex:10x:cellranger-mkref (data:seq:nucleotide: genome, data:annotation:gtf: annotation)[Source: v2.1.1]

Reference preparation tool for 10x Genomics Cell Ranger. Build a Cell Ranger-compatible reference from genome FASTA and gene GTF files. https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/references

genome

label: Reference genome
type: data:seq:nucleotide:
required: True
hidden: False

annotation

label: Annotation
type: data:annotation:gtf:
required: True
hidden: False

genome_index

label: Indexed genome
type: basic:dir
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

source

label: Gene ID source
type: basic:string
required: True
hidden: False

ChIP-Seq (Gene Score)¶

data:chipseq:genescorechipseq-genescore (data:chipseq:peakscore peakscore, basic:decimal fdr, basic:decimal pval, basic:decimal logratio)[Source: v1.2.1]

Chip-Seq analysis - Gene Score (BCM)

peakscore

label: PeakScore file
type: data:chipseq:peakscore
description: PeakScore file

fdr

label: FDR threshold
type: basic:decimal
description: FDR threshold value (default = 0.00005).
default: 5e-05

pval

label: Pval threshold
type: basic:decimal
description: Pval threshold value (default = 0.00005).
default: 5e-05

logratio

label: Log-ratio threshold
type: basic:decimal
description: Log-ratio threshold value (default = 2).
default: 2.0

genescore

label: Gene Score
type: basic:file

ChIP-Seq (Peak Score)¶

data:chipseq:peakscorechipseq-peakscore (data:chipseq:callpeak:macs2 peaks, data:bed bed)[Source: v2.2.1]

Chip-Seq analysis - Peak Score (BCM)

peaks

label: MACS2 results
type: data:chipseq:callpeak:macs2
description: MACS2 results file (NarrowPeak)

bed

label: BED file
type: data:bed

peak_score

label: Peak Score
type: basic:file

ChIP-seq (MACS2)¶

data:chipseq:batch:macs2macs2-batch (list:data:alignment:bam alignments, data:bed promoter, basic:boolean advanced, basic:boolean tagalign, basic:integer q_threshold, basic:integer n_sub, basic:boolean tn5, basic:integer shift, basic:string duplicates, basic:string duplicates_prepeak, basic:decimal qvalue, basic:decimal pvalue, basic:decimal pvalue_prepeak, basic:integer cap_num, basic:integer mfold_lower, basic:integer mfold_upper, basic:integer slocal, basic:integer llocal, basic:integer extsize, basic:integer shift, basic:integer band_width, basic:boolean nolambda, basic:boolean fix_bimodal, basic:boolean nomodel, basic:boolean nomodel_prepeak, basic:boolean down_sample, basic:boolean bedgraph, basic:boolean spmr, basic:boolean call_summits, basic:boolean broad, basic:decimal broad_cutoff, data:bed blacklist, basic:boolean calculate_enrichment, basic:integer profile_window, basic:string shift_size)[Source: v1.4.2]

This process runs MACS2 in batch mode. MACS2 analysis is triggered for pairs of samples as defined using treatment-background sample relations. If there are no sample relations defined, each sample is treated individually for the MACS analysis. Model-based Analysis of ChIP-Seq (MACS 2.0), is used to identify transcript factor binding sites. MACS 2.0 captures the influence of genome complexity to evaluate the significance of enriched ChIP regions, and MACS improves the spatial resolution of binding sites through combining the information of both sequencing tag position and orientation. It has also an option to link nearby peaks together in order to call broad peaks. See [here](https://github.com/taoliu/MACS/) for more information. In addition to peak-calling, this process computes ChIP-Seq and ATAC-Seq QC metrics. Process returns a QC metrics report, fragment length estimation, and a deduplicated tagAlign file. QC report contains ENCODE 3 proposed QC metrics – [NRF](https://www.encodeproject.org/data-standards/terms/), [PBC bottlenecking coefficients, NSC, and RSC](https://genome.ucsc.edu/ENCODE/qualityMetrics.html#chipSeq).

alignments

label: Aligned reads
type: list:data:alignment:bam
description: Select multiple treatment/background samples.

promoter

label: Promoter regions BED file
type: data:bed
description: BED file containing promoter regions (TSS+-1000 bp for example). Needed to get the number of peaks and reads mapped to promoter regions.
required: False

advanced

label: Show advanced options
type: basic:boolean
description: Inspect and modify parameters.
default: False

tagalign

label: Use tagAlign files
type: basic:boolean
description: Use filtered tagAlign files as case (treatment) and control (background) samples. If extsize parameter is not set, run MACS using input’s estimated fragment length.
hidden: !advanced
default: True

prepeakqc_settings.q_threshold

label: Quality filtering threshold
type: basic:integer
default: 30

prepeakqc_settings.n_sub

label: Number of reads to subsample
type: basic:integer
default: 15000000

prepeakqc_settings.tn5

label: Tn5 shifting
type: basic:boolean
description: Tn5 transposon shifting. Shift reads on “+” strand by 4 bp and reads on “-” strand by 5 bp.
default: False

prepeakqc_settings.shift

label: User-defined cross-correlation peak strandshift
type: basic:integer
description: If defined, SPP tool will not try to estimate fragment length but will use the given value as fragment length.
required: False

settings.duplicates

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

tagalign

choices

1: 1
auto: auto
all: all

settings.duplicates_prepeak

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

!tagalign

default

all

choices

1: 1
auto: auto
all: all

settings.qvalue

label: Q-value cutoff
type: basic:decimal
description: The q-value (minimum FDR) cutoff to call significant regions. Q-values are calculated from p-values using Benjamini-Hochberg procedure.
required: False
disabled: settings.pvalue && settings.pvalue_prepeak

settings.pvalue

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
required: False
disabled: settings.qvalue
hidden: tagalign

settings.pvalue_prepeak

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
disabled: settings.qvalue
hidden: !tagalign || settings.qvalue
default: 1e-05

settings.cap_num

label: Cap number of peaks by taking top N peaks
type: basic:integer
description: To keep all peaks set value to 0.
disabled: settings.broad
default: 500000

settings.mfold_lower

label: MFOLD range (lower limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.mfold_upper

label: MFOLD range (upper limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.slocal

label: Small local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.llocal

label: Large local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.extsize

label: extsize
type: basic:integer
description: While ‘–nomodel’ is set, MACS uses this parameter to extend reads in 5’->3’ direction to fix-sized fragments. For example, if the size of binding region for your transcription factor is 200 bp, and you want to bypass the model building by MACS, this parameter can be set as 200. This option is only valid when –nomodel is set or when MACS fails to build model and –fix-bimodal is on.
required: False

settings.shift

label: Shift
type: basic:integer
description: Note, this is NOT the legacy –shiftsize option which is replaced by –extsize! You can set an arbitrary shift in bp here. Please Use discretion while setting it other than default value (0). When –nomodel is set, MACS will use this value to move cutting ends (5’) then apply –extsize from 5’ to 3’ direction to extend them to fragments. When this value is negative, ends will be moved toward 3’->5’ direction, otherwise 5’->3’ direction. Recommended to keep it as default 0 for ChIP-Seq datasets, or -1 * half of EXTSIZE together with –extsize option for detecting enriched cutting loci such as certain DNAseI-Seq datasets. Note, you can’t set values other than 0 if format is BAMPE for paired-end data. Default is 0.
required: False

settings.band_width

label: Band width
type: basic:integer
description: The band width which is used to scan the genome ONLY for model building. You can set this parameter as the sonication fragment size expected from wet experiment. The previous side effect on the peak detection process has been removed. So this parameter only affects the model building.
required: False

settings.nolambda

label: Use backgroud lambda as local lambda
type: basic:boolean
description: With this flag on, MACS will use the background lambda as local lambda. This means MACS will not consider the local bias at peak candidate regions.
default: False

settings.fix_bimodal

label: Turn on the auto paired-peak model process
type: basic:boolean
description: Turn on the auto paired-peak model process. If it’s set, when MACS failed to build paired model, it will use the nomodel settings, the ‘–extsize’ parameter to extend each tag. If set, MACS will be terminated if paired-peak model has failed.
default: False

settings.nomodel

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: tagalign
default: False

settings.nomodel_prepeak

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: !tagalign
default: True

settings.down_sample

label: Down-sample
type: basic:boolean
description: When set to true, random sampling method will scale down the bigger sample. By default, MACS uses linear scaling. This option will make the results unstable and irreproducible since each time, random reads would be selected, especially the numbers (pileup, pvalue, qvalue) would change.
default: False

settings.bedgraph

label: Save fragment pileup and control lambda
type: basic:boolean
description: If this flag is on, MACS will store the fragment pileup, control lambda, -log10pvalue and -log10qvalue scores in bedGraph files. The bedGraph files will be stored in current directory named NAME+’_treat_pileup.bdg’ for treatment data, NAME+’_control_lambda.bdg’ for local lambda values from control, NAME+’_treat_pvalue.bdg’ for Poisson pvalue scores (in -log10(pvalue) form), and NAME+’_treat_qvalue.bdg’ for q-value scores from Benjamini-Hochberg-Yekutieli procedure.
default: True

settings.spmr

label: Save signal per million reads for fragment pileup profiles
type: basic:boolean
disabled: settings.bedgraph === false
default: True

settings.call_summits

label: Call summits
type: basic:boolean
description: MACS will now reanalyze the shape of signal profile (p or q-score depending on cutoff setting) to deconvolve subpeaks within each peak called from general procedure. It’s highly recommended to detect adjacent binding events. While used, the output subpeaks of a big peak region will have the same peak boundaries, and different scores and peak summit positions.
default: False

settings.broad

label: Composite broad regions
type: basic:boolean
description: When this flag is on, MACS will try to composite broad regions in BED12 (a gene-model-like format) by putting nearby highly enriched regions into a broad region with loose cutoff. The broad region is controlled by another cutoff through –broad-cutoff. The maximum length of broad region length is 4 times of d from MACS.
disabled: settings.call_summits === true
default: False

settings.broad_cutoff

label: Broad cutoff
type: basic:decimal
description: Cutoff for broad region. This option is not available unless –broad is set. If -p is set, this is a p-value cutoff, otherwise, it’s a q-value cutoff. DEFAULT = 0.1
required: False
disabled: settings.call_summits === true || settings.broad !== true

chipqc_settings.blacklist

label: Blacklist regions
type: data:bed
description: BED file containing genomic regions that should be excluded from the analysis.
required: False

chipqc_settings.calculate_enrichment

label: Calculate enrichment
type: basic:boolean
description: Calculate enrichment of signal in known genomic annotation. By default annotation is provided from the TranscriptDB package specified by genome bulid which should match one of the supported annotations (hg19, hg38, hg18, mm10, mm9, rn4, ce6, dm3). If annotation is not supported the analysis is skipped.
default: False

chipqc_settings.profile_window

label: Window size
type: basic:integer
description: An integer indicating the width of the window used for peak profiles. Peaks will be centered on their summits and include half of the window size upstream and half downstream of this point.
default: 400

chipqc_settings.shift_size

label: Shift size
type: basic:string
description: Vector of values to try when computing optimal shift sizes. It should be specifeird as consecutive numbers vector with start:end
default: 1:300

ChIP-seq (MACS2-ROSE2)¶

data:chipseq:batch:macs2macs2-rose2-batch (list:data:alignment:bam alignments, data:bed promoter, basic:boolean advanced, basic:boolean tagalign, basic:integer q_threshold, basic:integer n_sub, basic:boolean tn5, basic:integer shift, basic:string duplicates, basic:string duplicates_prepeak, basic:decimal qvalue, basic:decimal pvalue, basic:decimal pvalue_prepeak, basic:integer cap_num, basic:integer mfold_lower, basic:integer mfold_upper, basic:integer slocal, basic:integer llocal, basic:integer extsize, basic:integer shift, basic:integer band_width, basic:boolean nolambda, basic:boolean fix_bimodal, basic:boolean nomodel, basic:boolean nomodel_prepeak, basic:boolean down_sample, basic:boolean bedgraph, basic:boolean spmr, basic:boolean call_summits, basic:boolean broad, basic:decimal broad_cutoff, basic:boolean use_filtered_bam, basic:integer tss, basic:integer stitch, data:bed mask, data:bed blacklist, basic:boolean calculate_enrichment, basic:integer profile_window, basic:string shift_size)[Source: v1.4.2]

This process runs MACS2 in batch mode. MACS2 analysis is triggered for pairs of samples as defined using treatment-background sample relations. If there are no sample relations defined, each sample is treated individually for the MACS analysis. Model-based Analysis of ChIP-Seq (MACS 2.0), is used to identify transcript factor binding sites. MACS 2.0 captures the influence of genome complexity to evaluate the significance of enriched ChIP regions, and MACS improves the spatial resolution of binding sites through combining the information of both sequencing tag position and orientation. It has also an option to link nearby peaks together in order to call broad peaks. See [here](https://github.com/taoliu/MACS/) for more information. In addition to peak-calling, this process computes ChIP-Seq and ATAC-Seq QC metrics. Process returns a QC metrics report, fragment length estimation, and a deduplicated tagAlign file. QC report contains ENCODE 3 proposed QC metrics – [NRF](https://www.encodeproject.org/data-standards/terms/), [PBC bottlenecking coefficients, NSC, and RSC](https://genome.ucsc.edu/ENCODE/qualityMetrics.html#chipSeq). For identification of super enhancers R2 uses the Rank Ordering of Super-Enhancers algorithm (ROSE2). This takes the peaks called by RSEG for acetylation and calculates the distances in-between to judge whether they can be considered super-enhancers. The ranked values can be plotted and by locating the inflection point in the resulting graph, super-enhancers can be assigned. It can also be used with the MACS calculated data. See [here](http://younglab.wi.mit.edu/super_enhancer_code.html) for more information.

alignments

label: Aligned reads
type: list:data:alignment:bam
description: Select multiple treatment/background samples.

promoter

label: Promoter regions BED file
type: data:bed
description: BED file containing promoter regions (TSS+-1000 bp for example). Needed to get the number of peaks and reads mapped to promoter regions.
required: False

advanced

label: Show advanced options
type: basic:boolean
description: Inspect and modify parameters.
default: False

tagalign

label: Use tagAlign files
type: basic:boolean
description: Use filtered tagAlign files as case (treatment) and control (background) samples. If extsize parameter is not set, run MACS using input’s estimated fragment length.
hidden: !advanced
default: True

prepeakqc_settings.q_threshold

label: Quality filtering threshold
type: basic:integer
default: 30

prepeakqc_settings.n_sub

label: Number of reads to subsample
type: basic:integer
default: 15000000

prepeakqc_settings.tn5

label: Tn5 shifting
type: basic:boolean
description: Tn5 transposon shifting. Shift reads on “+” strand by 4 bp and reads on “-” strand by 5 bp.
default: False

prepeakqc_settings.shift

label: User-defined cross-correlation peak strandshift
type: basic:integer
description: If defined, SPP tool will not try to estimate fragment length but will use the given value as fragment length.
required: False

settings.duplicates

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

tagalign

choices

1: 1
auto: auto
all: all

settings.duplicates_prepeak

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

required

False

hidden

!tagalign

default

all

choices

1: 1
auto: auto
all: all

settings.qvalue

label: Q-value cutoff
type: basic:decimal
description: The q-value (minimum FDR) cutoff to call significant regions. Q-values are calculated from p-values using Benjamini-Hochberg procedure.
required: False
disabled: settings.pvalue && settings.pvalue_prepeak

settings.pvalue

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
required: False
disabled: settings.qvalue
hidden: tagalign

settings.pvalue_prepeak

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff. If specified, MACS2 will use p-value instead of q-value cutoff.
disabled: settings.qvalue
hidden: !tagalign || settings.qvalue
default: 1e-05

settings.cap_num

label: Cap number of peaks by taking top N peaks
type: basic:integer
description: To keep all peaks set value to 0.
disabled: settings.broad
default: 500000

settings.mfold_lower

label: MFOLD range (lower limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.mfold_upper

label: MFOLD range (upper limit)
type: basic:integer
description: This parameter is used to select the regions within MFOLD range of high-confidence enrichment ratio against background to build model. The regions must be lower than upper limit, and higher than the lower limit of fold enrichment. DEFAULT:10,30 means using all regions not too low (>10) and not too high (<30) to build paired-peaks model. If MACS can not find more than 100 regions to build model, it will use the –extsize parameter to continue the peak detection ONLY if –fix-bimodal is set.
required: False

settings.slocal

label: Small local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.llocal

label: Large local region
type: basic:integer
description: Slocal and llocal parameters control which two levels of regions will be checked around the peak regions to calculate the maximum lambda as local lambda. By default, MACS considers 1000 bp for small local region (–slocal), and 10000 bp for large local region (–llocal) which captures the bias from a long range effect like an open chromatin domain. You can tweak these according to your project. Remember that if the region is set too small, a sharp spike in the input data may kill the significant peak.
required: False

settings.extsize

label: extsize
type: basic:integer
description: While ‘–nomodel’ is set, MACS uses this parameter to extend reads in 5’->3’ direction to fix-sized fragments. For example, if the size of binding region for your transcription factor is 200 bp, and you want to bypass the model building by MACS, this parameter can be set as 200. This option is only valid when –nomodel is set or when MACS fails to build model and –fix-bimodal is on.
required: False

settings.shift

label: Shift
type: basic:integer
description: Note, this is NOT the legacy –shiftsize option which is replaced by –extsize! You can set an arbitrary shift in bp here. Please Use discretion while setting it other than default value (0). When –nomodel is set, MACS will use this value to move cutting ends (5’) then apply –extsize from 5’ to 3’ direction to extend them to fragments. When this value is negative, ends will be moved toward 3’->5’ direction, otherwise 5’->3’ direction. Recommended to keep it as default 0 for ChIP-Seq datasets, or -1 * half of EXTSIZE together with –extsize option for detecting enriched cutting loci such as certain DNAseI-Seq datasets. Note, you can’t set values other than 0 if format is BAMPE for paired-end data. Default is 0.
required: False

settings.band_width

label: Band width
type: basic:integer
description: The band width which is used to scan the genome ONLY for model building. You can set this parameter as the sonication fragment size expected from wet experiment. The previous side effect on the peak detection process has been removed. So this parameter only affects the model building.
required: False

settings.nolambda

label: Use backgroud lambda as local lambda
type: basic:boolean
description: With this flag on, MACS will use the background lambda as local lambda. This means MACS will not consider the local bias at peak candidate regions.
default: False

settings.fix_bimodal

label: Turn on the auto paired-peak model process
type: basic:boolean
description: Turn on the auto paired-peak model process. If it’s set, when MACS failed to build paired model, it will use the nomodel settings, the ‘–extsize’ parameter to extend each tag. If set, MACS will be terminated if paired-peak model has failed.
default: False

settings.nomodel

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: tagalign
default: False

settings.nomodel_prepeak

label: Bypass building the shifting model
type: basic:boolean
description: While on, MACS will bypass building the shifting model.
hidden: !tagalign
default: True

settings.down_sample

label: Down-sample
type: basic:boolean
description: When set to true, random sampling method will scale down the bigger sample. By default, MACS uses linear scaling. This option will make the results unstable and irreproducible since each time, random reads would be selected, especially the numbers (pileup, pvalue, qvalue) would change.
default: False

settings.bedgraph

label: Save fragment pileup and control lambda
type: basic:boolean
description: If this flag is on, MACS will store the fragment pileup, control lambda, -log10pvalue and -log10qvalue scores in bedGraph files. The bedGraph files will be stored in current directory named NAME+’_treat_pileup.bdg’ for treatment data, NAME+’_control_lambda.bdg’ for local lambda values from control, NAME+’_treat_pvalue.bdg’ for Poisson pvalue scores (in -log10(pvalue) form), and NAME+’_treat_qvalue.bdg’ for q-value scores from Benjamini-Hochberg-Yekutieli procedure.
default: True

settings.spmr

label: Save signal per million reads for fragment pileup profiles
type: basic:boolean
disabled: settings.bedgraph === false
default: True

settings.call_summits

label: Call summits
type: basic:boolean
description: MACS will now reanalyze the shape of signal profile (p or q-score depending on cutoff setting) to deconvolve subpeaks within each peak called from general procedure. It’s highly recommended to detect adjacent binding events. While used, the output subpeaks of a big peak region will have the same peak boundaries, and different scores and peak summit positions.
default: False

settings.broad

label: Composite broad regions
type: basic:boolean
description: When this flag is on, MACS will try to composite broad regions in BED12 (a gene-model-like format) by putting nearby highly enriched regions into a broad region with loose cutoff. The broad region is controlled by another cutoff through –broad-cutoff. The maximum length of broad region length is 4 times of d from MACS.
disabled: settings.call_summits === true
default: False

settings.broad_cutoff

label: Broad cutoff
type: basic:decimal
description: Cutoff for broad region. This option is not available unless –broad is set. If -p is set, this is a p-value cutoff, otherwise, it’s a q-value cutoff. DEFAULT = 0.1
required: False
disabled: settings.call_summits === true || settings.broad !== true

rose_settings.use_filtered_bam

label: Use Filtered BAM File
type: basic:boolean
description: Use filtered BAM file from a MACS2 object to rank enhancers by.
default: True

rose_settings.tss

label: TSS exclusion
type: basic:integer
description: Enter a distance from TSS to exclude. 0 = no TSS exclusion
default: 0

rose_settings.stitch

label: Stitch
type: basic:integer
description: Enter a max linking distance for stitching. If not given, optimal stitching parameter will be determined automatically.
required: False

rose_settings.mask

label: Masking BED file
type: data:bed
description: Mask a set of regions from analysis. Provide a BED of masking regions.
required: False

chipqc_settings.blacklist

label: Blacklist regions
type: data:bed
description: BED file containing genomic regions that should be excluded from the analysis.
required: False

chipqc_settings.calculate_enrichment

label: Calculate enrichment
type: basic:boolean
description: Calculate enrichment of signal in known genomic annotation. By default annotation is provided from the TranscriptDB package specified by genome bulid which should match one of the supported annotations (hg19, hg38, hg18, mm10, mm9, rn4, ce6, dm3). If annotation is not supported the analysis is skipped.
default: False

chipqc_settings.profile_window

label: Window size
type: basic:integer
description: An integer indicating the width of the window used for peak profiles. Peaks will be centered on their summits and include half of the window size upstream and half downstream of this point.
default: 400

chipqc_settings.shift_size

label: Shift size
type: basic:string
description: Vector of values to try when computing optimal shift sizes. It should be specifeird as consecutive numbers vector with start:end
default: 1:300

Chemical Mutagenesis¶

data:workflow:chemutworkflow-chemut (basic:string analysis_type, data:seq:nucleotide genome, list:data:alignment:bam parental_strains, list:data:alignment:bam mutant_strains, basic:boolean advanced, basic:boolean br_and_ind_ra, basic:boolean dbsnp, data:variants:vcf known_sites, list:data:variants:vcf known_indels, basic:integer stand_emit_conf, basic:integer stand_call_conf, basic:boolean rf, basic:boolean advanced, basic:integer read_depth)[Source: v1.0.2]

analysis_type

label

Analysis type

type

basic:string

description

Choice of the analysis type. Use “SNV” or “INDEL” options to run the GATK analysis only on the haploid portion of the dicty genome. Choose options SNV_CHR2 or INDEL_CHR2 to run the analysis only on the diploid portion of CHR2 (-ploidy 2 -L chr2:2263132-3015703).

default

snv

choices

SNV: snv
INDEL: indel
SNV_CHR2: snv_chr2
INDEL_CHR2: indel_chr2

genome

label: Reference genome
type: data:seq:nucleotide

parental_strains

label: Parental strains
type: list:data:alignment:bam

mutant_strains

label: Mutant strains
type: list:data:alignment:bam

Vc.advanced

label: Advanced options
type: basic:boolean
required: False
default: False

Vc.br_and_ind_ra

label: Do variant base recalibration and indel realignment
type: basic:boolean
required: False
hidden: Vc.advanced === false
default: False

Vc.dbsnp

label: Use dbSNP file
type: basic:boolean
description: rsIDs from this file are used to populate the ID column of the output. Also, the DB INFO flag will be set when appropriate. dbSNP is not used in any way for the calculations themselves.
required: False
hidden: Vc.advanced === false
default: False

Vc.known_sites

label: Known sites (dbSNP)
type: data:variants:vcf
required: False
hidden: Vc.advanced === false || Vc.br_and_ind_ra === false && Vc.dbsnp === false

Vc.known_indels

label: Known indels
type: list:data:variants:vcf
required: False
hidden: Vc.advanced === false || Vc.br_and_ind_ra === false

Vc.stand_emit_conf

label: Emission confidence threshold
type: basic:integer
description: The minimum confidence threshold (phred-scaled) at which the program should emit sites that appear to be possibly variant.
required: False
hidden: Vc.advanced === false
default: 10

Vc.stand_call_conf

label: Calling confidence threshold
type: basic:integer
description: The minimum confidence threshold (phred-scaled) at which the program should emit variant sites as called. If a site’s associated genotype has a confidence score lower than the calling threshold, the program will emit the site as filtered and will annotate it as LowQual. This threshold separates high confidence calls from low confidence calls.
required: False
hidden: Vc.advanced === false
default: 30

Vc.rf

label: ReasignOneMappingQuality Filter
type: basic:boolean
description: This read transformer will change a certain read mapping quality to a different value without affecting reads that have other mapping qualities. This is intended primarily for users of RNA-Seq data handling programs such as TopHat, which use MAPQ = 255 to designate uniquely aligned reads. According to convention, 255 normally designates “unknown” quality, and most GATK tools automatically ignore such reads. By reassigning a different mapping quality to those specific reads, users of TopHat and other tools can circumvent this problem without affecting the rest of their dataset.
required: False
hidden: Vc.advanced === false
default: False

Vf.advanced

label: Advanced options
type: basic:boolean
required: False
default: False

Vf.read_depth

label: Read depth cutoff
type: basic:integer
description: The minimum number of replicate reads required for a variant site to be included.
required: False
hidden: Vf.advanced === false
default: 5

ChipQC¶

data:chipqc:chipqc (data:alignment:bam alignment, data:chipseq:callpeak peaks, data:bed blacklist, basic:boolean calculate_enrichment, basic:integer quality_threshold, basic:integer profile_window, basic:string shift_size)[Source: v1.1.1]

Calculate quality control metrics for ChIP-seq samples. The analysis is based on ChIPQC package which computs a variety of quality control metrics and statistics, and provides plots and a report for assessment of experimental data for further analysis.

alignment

label: Aligned reads
type: data:alignment:bam
required: True
hidden: False

peaks

label: Called peaks
type: data:chipseq:callpeak
required: True
hidden: False

blacklist

label: Blacklist regions
type: data:bed
description: BED file containing genomic regions that should be excluded from the analysis.
required: False
hidden: False

calculate_enrichment

label: Calculate enrichment
type: basic:boolean
description: Calculate enrichment of signal in known genomic annotation. By default annotation is provided from the TranscriptDB package specified by genome bulid which should match one of the supported annotations (hg19, hg38, hg18, mm10, mm9, rn4, ce6, dm3). If annotation is not supported the analysis is skipped.
required: True
hidden: False
default: False

advanced.quality_threshold

label: Mapping quality threshold
type: basic:integer
description: Only reads with mapping quality scores above this threshold will be used for some statistics.
required: True
hidden: False
default: 15

advanced.profile_window

label: Window size
type: basic:integer
description: An integer indicating the width of the window used for peak profiles. Peaks will be centered on their summits and include half of the window size upstream and half downstream of this point.
required: True
hidden: False
default: 400

advanced.shift_size

label: Shift size
type: basic:string
description: Vector of values to try when computing optimal shift sizes. It should be specifeird as consecutive numbers vector with start:end
required: True
hidden: False
default: 1:300

report_folder

label: ChipQC report folder
type: basic:dir
required: True
hidden: False

ccplot

label: Cross coverage score plot
type: basic:file
required: True
hidden: False

coverage_histogram

label: SSD metric plot
type: basic:file
required: True
hidden: False

peak_profile

label: Peak profile plot
type: basic:file
required: True
hidden: False

peaks_barplot

label: Barplot of reads in peaks
type: basic:file
required: True
hidden: False

peaks_density_plot

label: Density plot of reads in peaks
type: basic:file
required: True
hidden: False

enrichment_heatmap

label: Heatmap of reads in genomic features
type: basic:file
required: False
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Convert GFF3 to GTF¶

data:annotation:gtfgff-to-gtf (data:annotation:gff3 annotation)[Source: v0.5.1]

Convert GFF3 file to GTF format.

annotation

label: Annotation (GFF3)
type: data:annotation:gff3
description: Annotation in GFF3 format.

annot

label: Converted GTF file
type: basic:file

annot_sorted

label: Sorted GTF file
type: basic:file

annot_sorted_idx_igv

label: Igv index for sorted GTF file
type: basic:file

annot_sorted_track_jbrowse

label: Jbrowse track for sorted GTF
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Convert files to reads (paired-end)¶

data:reads:fastq:pairedfiles-to-fastq-paired (list:data:file src1, list:data:file src2, basic:boolean merge_lanes)[Source: v1.4.1]

Convert FASTQ files to paired-end reads.

src1

label: Mate1
type: list:data:file

src2

label: Mate2
type: list:data:file

merge_lanes

label: Merge lanes
type: basic:boolean
description: Merge paired-end sample data split into multiple sequencing lanes into a single pair of FASTQ files.
default: False

fastq

label: Reads file (mate 1)
type: list:basic:file

fastq2

label: Reads file (mate 2)
type: list:basic:file

fastqc_url

label: Quality control with FastQC (Upstream)
type: list:basic:file:html

fastqc_url2

label: Quality control with FastQC (Downstream)
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive (Upstream)
type: list:basic:file

fastqc_archive2

label: Download FastQC archive (Downstream)
type: list:basic:file

Convert files to reads (single-end)¶

data:reads:fastq:singlefiles-to-fastq-single (list:data:file src, basic:boolean merge_lanes)[Source: v1.4.1]

Convert FASTQ files to single-end reads.

src

label: Reads
type: list:data:file
description: Sequencing reads in FASTQ format

merge_lanes

label: Merge lanes
type: basic:boolean
description: Merge sample data split into multiple sequencing lanes into a single FASTQ file.
default: False

fastq

label: Reads file
type: list:basic:file

fastqc_url

label: Quality control with FastQC
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive
type: list:basic:file

Cuffdiff 2.2¶

data:differentialexpression:cuffdiff:cuffdiff (list:data:cufflinks:cuffquant case, list:data:cufflinks:cuffquant control, list:basic:string labels, data:annotation annotation, data:seq:nucleotide genome, basic:boolean multi_read_correct, basic:boolean create_sets, basic:decimal gene_logfc, basic:decimal gene_fdr, basic:decimal fdr, basic:string library_type, basic:string library_normalization, basic:string dispersion_method)[Source: v3.3.2]

Run Cuffdiff 2.2 analysis. Cuffdiff finds significant changes in transcript expression, splicing, and promoter use. You can use it to find differentially expressed genes and transcripts, as well as genes that are being differentially regulated at the transcriptional and post-transcriptional level. See [here](http://cole-trapnell-lab.github.io/cufflinks/cuffdiff/) and [here](https://software.broadinstitute.org/cancer/software/genepattern/modules/docs/Cuffdiff/7) for more information.

case

label: Case samples
type: list:data:cufflinks:cuffquant
required: True
hidden: False

control

label: Control samples
type: list:data:cufflinks:cuffquant
required: True
hidden: False

labels

label: Group labels
type: list:basic:string
description: Define labels for each sample group.
required: True
hidden: False
default: ['control', 'case']

annotation

label: Annotation (GTF/GFF3)
type: data:annotation
description: A transcript annotation file produced by cufflinks, cuffcompare, or other tool.
required: True
hidden: False

genome

label: Run bias detection and correction algorithm
type: data:seq:nucleotide
description: Provide Cufflinks with a multifasta file (genome file) via this option to instruct it to run a bias detection and correction algorithm which can significantly improve accuracy of transcript abundance estimates.
required: False
hidden: False

multi_read_correct

label: Do initial estimation procedure to more accurately weight reads with multiple genome mappings
type: basic:boolean
required: True
hidden: False
default: False

create_sets

label: Create gene sets
type: basic:boolean
description: After calculating differential gene expressions create gene sets for up-regulated genes, down-regulated genes and all genes.
required: True
hidden: False
default: False

gene_logfc

label: Log2 fold change threshold for gene sets
type: basic:decimal
description: Genes above Log2FC are considered as up-regulated and genes below -Log2FC as down-regulated.
required: True
hidden: !create_sets
default: 1.0

gene_fdr

label: FDR threshold for gene sets
type: basic:decimal
required: True
hidden: !create_sets
default: 0.05

fdr

label: Allowed FDR
type: basic:decimal
description: The allowed false discovery rate. The default is 0.05.
required: True
hidden: False
default: 0.05

library_type

label

Library type

type

basic:string

description

In cases where Cufflinks cannot determine the platform and protocol used to generate input reads, you can supply this information manually, which will allow Cufflinks to infer source strand information with certain protocols. The available options are listed below. For paired-end data, we currently only support protocols where reads point towards each other: fr-unstranded - Reads from the left-most end of the fragment (in transcript coordinates) map to the transcript strand and the right-most end maps to the opposite strand; fr-firststrand - Same as above except we enforce the rule that the right-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during first strand synthesis is sequenced; fr-secondstrand - Same as above except we enforce the rule that the left-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during second strand synthesis is sequenced.

required

True

hidden

False

default

fr-unstranded

choices

fr-unstranded: fr-unstranded
fr-firststrand: fr-firststrand
fr-secondstrand: fr-secondstrand

library_normalization

label

Library normalization method

type

basic:string

description

You can control how library sizes (i.e. sequencing depths) are normalized in Cufflinks and Cuffdiff. Cuffdiff has several methods that require multiple libraries in order to work. Library normalization methods supported by Cufflinks work on one library at a time.

required

True

hidden

False

default

geometric

choices

geometric: geometric
classic-fpkm: classic-fpkm
quartile: quartile

dispersion_method

label

Dispersion method

type

basic:string

description

Cuffdiff works by modeling the variance in fragment counts across replicates as a function of the mean fragment count across replicates. Strictly speaking, models a quantitity called dispersion - the variance present in a group of samples beyond what is expected from a simple Poisson model of RNA_Seq. You can control how Cuffdiff constructs its model of dispersion in locus fragment counts. Each condition that has replicates can receive its own model, or Cuffdiff can use a global model for all conditions. All of these policies are identical to those used by DESeq (Anders and Huber, Genome Biology, 2010).

required

True

hidden

False

default

pooled

choices

pooled: pooled
per-condition: per-condition
blind: blind
poisson: poisson

raw

label: Differential expression
type: basic:file
required: True
hidden: False

de_json

label: Results table (JSON)
type: basic:json
required: True
hidden: False

de_file

label: Results table (file)
type: basic:file
required: True
hidden: False

transcript_diff_exp

label: Differential expression (transcript level)
type: basic:file
required: True
hidden: False

tss_group_diff_exp

label: Differential expression (primary transcript)
type: basic:file
required: True
hidden: False

cds_diff_exp

label: Differential expression (coding sequence)
type: basic:file
required: True
hidden: False

cuffdiff_output

label: Cuffdiff output
type: basic:file
required: True
hidden: False

source

label: Gene ID database
type: basic:string
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

feature_type

label: Feature type
type: basic:string
required: True
hidden: False

Cufflinks 2.2¶

data:cufflinks:cufflinkscufflinks (data:alignment:bam alignment, data:annotation annotation, data:seq:nucleotide genome, data:annotation:gtf mask_file, basic:string library_type, basic:string annotation_usage, basic:boolean multi_read_correct)[Source: v3.1.1]

Cufflinks assembles transcripts, estimates their abundances, and tests for differential expression and regulation in RNA-Seq samples. It accepts aligned RNA-Seq reads and assembles the alignments into a parsimonious set of transcripts. Cufflinks then estimates the relative abundances of these transcripts based on how many reads support each one, taking into account biases in library preparation protocols. See [here](http://cole-trapnell-lab.github.io/cufflinks/) for more information.

alignment

label: Aligned reads
type: data:alignment:bam

annotation

label: Annotation (GTF/GFF3)
type: data:annotation
required: False

genome

label: Run bias detection and correction algorithm
type: data:seq:nucleotide
description: Provide Cufflinks with a multifasta file (genome file) via this option to instruct it to run a bias detection and correction algorithm which can significantly improve accuracy of transcript abundance estimates.
required: False

mask_file

label: Mask file
type: data:annotation:gtf
description: Ignore all reads that could have come from transcripts in this GTF file. We recommend including any annotated rRNA, mitochondrial transcripts other abundant transcripts you wish to ignore in your analysis in this file. Due to variable efficiency of mRNA enrichment methods and rRNA depletion kits, masking these transcripts often improves the overall robustness of transcript abundance estimates.
required: False

library_type

label

Library type

type

basic:string

description

In cases where Cufflinks cannot determine the platform and protocol used to generate input reads, you can supply this information manually, which will allow Cufflinks to infer source strand information with certain protocols. The available options are listed below. For paired-end data, we currently only support protocols where reads are point towards each other: fr-unstranded - Reads from the left-most end of the fragment (in transcript coordinates) map to the transcript strand, and the right-most end maps to the opposite strand; fr-firststrand - Same as above except we enforce the rule that the right-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during first strand synthesis is sequenced; fr-secondstrand - Same as above except we enforce the rule that the left-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during second strand synthesis is sequenced.

default

fr-unstranded

choices

fr-unstranded: fr-unstranded
fr-firststrand: fr-firststrand
fr-secondstrand: fr-secondstrand

annotation_usage

label

Instruct Cufflinks how to use the provided annotation (GFF/GTF) file

type

basic:string

description

GTF-guide - tells Cufflinks to use the supplied reference annotation (GFF) to guide RABT assembly. Reference transcripts will be tiled with faux-reads to provide additional information in assembly. Output will include all reference transcripts as well as any novel genes and isoforms that are assembled. –GTF - tells Cufflinks to use the supplied reference annotation (a GFF file) to estimate isoform expression. It will not assemble novel transcripts, and the program will ignore alignments not structurally compatible with any reference transcript.

default

--GTF-guide

choices

Use supplied reference annotation to guide RABT assembly (–GTF-guide): --GTF-guide
Use supplied reference annotation to estimate isoform expression (–GTF): --GTF

multi_read_correct

label: Do initial estimation procedure to more accurately weight reads with multiple genome mappings
type: basic:boolean
description: Run an initial estimation procedure that weights reads mapping to multiple locations more accurately.
default: False

transcripts

label: Assembled transcript isoforms
type: basic:file

isoforms_fpkm_tracking

label: Isoforms FPKM tracking
type: basic:file

genes_fpkm_tracking

label: Genes FPKM tracking
type: basic:file

skipped_loci

label: Skipped loci
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Cuffmerge¶

data:annotation:cuffmergecuffmerge (list:data:cufflinks:cufflinks expressions, list:data:annotation:gtf gtf, data:annotation gff, data:seq:nucleotide genome, basic:integer threads)[Source: v2.1.1]

Cufflinks includes a script called Cuffmerge that you can use to merge together several Cufflinks assemblies. It also handles running Cuffcompare for you, and automatically filters a number of transfrags that are probably artfifacts. The main purpose of Cuffmerge is to make it easier to make an assembly GTF file suitable for use with Cuffdiff. See [here](http://cole-trapnell-lab.github.io/cufflinks/cuffmerge/) for more information.

expressions

label: Cufflinks transcripts (GTF)
type: list:data:cufflinks:cufflinks
required: False

gtf

label: Annotation files (GTF)
type: list:data:annotation:gtf
description: Annotation files you wish to merge together with Cufflinks produced annotation files (e.g. upload Cufflinks annotation GTF file)
required: False

gff

label: Reference annotation (GTF/GFF3)
type: data:annotation
description: An optional “reference” annotation GTF. The input assemblies are merged together with the reference GTF and included in the final output.
required: False

genome

label: Reference genome
type: data:seq:nucleotide
description: This argument should point to the genomic DNA sequences for the reference. If a directory, it should contain one fasta file per contig. If a multifasta file, all contigs should be present. The merge script will pass this option to cuffcompare, which will use the sequences to assist in classifying transfrags and excluding artifacts (e.g. repeats). For example, Cufflinks transcripts consisting mostly of lower-case bases are classified as repeats. Note that <seq_dir> must contain one fasta file per reference chromosome, and each file must be named after the chromosome, and have a .fa or .fasta extension
required: False

threads

label: Use this many processor threads
type: basic:integer
description: Use this many threads to align reads. The default is 1.
default: 1

annot

label: Merged GTF file
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Cuffnorm¶

data:cuffnormcuffnorm (list:data:cufflinks:cuffquant cuffquant, data:annotation annotation, basic:boolean useERCC)[Source: v2.3.1]

Cufflinks includes a program, Cuffnorm, that you can use to generate tables of expression values that are properly normalized for library size. Cuffnorm takes a GTF2/GFF3 file of transcripts as input, along with two or more SAM, BAM, or CXB files for two or more samples. See [here](http://cole-trapnell-lab.github.io/cufflinks/cuffnorm/) for more information. Replicate relation needs to be defined for Cuffnorm to account for replicates. If the replicate relation is not defined, each sample will be treated individually.

cuffquant

label: Cuffquant expression file
type: list:data:cufflinks:cuffquant

annotation

label: Annotation (GTF/GFF3)
type: data:annotation
description: A transcript annotation file produced by cufflinks, cuffcompare, or other source.

useERCC

label: ERCC spike-in normalization
type: basic:boolean
description: Use ERRCC spike-in controls for normalization.
default: False

genes_count

label: Genes count
type: basic:file

genes_fpkm

label: Genes FPKM
type: basic:file

genes_attr

label: Genes attr table
type: basic:file

isoform_count

label: Isoform count
type: basic:file

isoform_fpkm

label: Isoform FPKM
type: basic:file

isoform_attr

label: Isoform attr table
type: basic:file

cds_count

label: CDS count
type: basic:file

cds_fpkm

label: CDS FPKM
type: basic:file

cds_attr

label: CDS attr table
type: basic:file

tss_groups_count

label: TSS groups count
type: basic:file

tss_groups_fpkm

label: TSS groups FPKM
type: basic:file

tss_attr

label: TSS attr table
type: basic:file

run_info

label: Run info
type: basic:file

raw_scatter

label: FPKM exp scatter plot
type: basic:file

boxplot

label: Boxplot
type: basic:file

fpkm_exp_raw

label: FPKM exp raw
type: basic:file

replicate_correlations

label: Replicate correlatios plot
type: basic:file

fpkm_means

label: FPKM means
type: basic:file

exp_fpkm_means

label: Exp FPKM means
type: basic:file

norm_scatter

label: FKPM exp scatter normalized plot
type: basic:file
required: False

fpkm_exp_norm

label: FPKM exp normalized
type: basic:file
required: False

spike_raw

label: Spike raw
type: basic:file
required: False

spike_norm

label: Spike normalized
type: basic:file
required: False

R_data

label: All R normalization data
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Cuffquant 2.2¶

data:cufflinks:cuffquantcuffquant (data:alignment:bam alignment, data:annotation annotation, data:seq:nucleotide genome, data:annotation:gtf mask_file, basic:string library_type, basic:boolean multi_read_correct)[Source: v2.1.1]

Cuffquant allows you to compute the gene and transcript expression profiles and save these profiles to files that you can analyze later with Cuffdiff or Cuffnorm. See [here](http://cole-trapnell-lab.github.io/cufflinks/manual/) for more information.

alignment

label: Aligned reads
type: data:alignment:bam

annotation

label: Annotation (GTF/GFF3)
type: data:annotation

genome

label: Run bias detection and correction algorithm
type: data:seq:nucleotide
description: Provide Cufflinks with a multifasta file (genome file) via this option to instruct it to run a bias detection and correction algorithm which can significantly improve accuracy of transcript abundance estimates.
required: False

mask_file

label: Mask file
type: data:annotation:gtf
description: Ignore all reads that could have come from transcripts in this GTF file. We recommend including any annotated rRNA, mitochondrial transcripts other abundant transcripts you wish to ignore in your analysis in this file. Due to variable efficiency of mRNA enrichment methods and rRNA depletion kits, masking these transcripts often improves the overall robustness of transcript abundance estimates.
required: False

library_type

label

Library type

type

basic:string

description

In cases where Cufflinks cannot determine the platform and protocol used to generate input reads, you can supply this information manually, which will allow Cufflinks to infer source strand information with certain protocols. The available options are listed below. For paired-end data, we currently only support protocols where reads are point towards each other: fr-unstranded - Reads from the left-most end of the fragment (in transcript coordinates) map to the transcript strand, and the right-most end maps to the opposite strand; fr-firststrand - Same as above except we enforce the rule that the right-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during first strand synthesis is sequenced; fr-secondstrand - Same as above except we enforce the rule that the left-most end of the fragment (in transcript coordinates) is the first sequenced (or only sequenced for single-end reads). Equivalently, it is assumed that only the strand generated during second strand synthesis is sequenced.

default

fr-unstranded

choices

fr-unstranded: fr-unstranded
fr-firststrand: fr-firststrand
fr-secondstrand: fr-secondstrand

multi_read_correct

label: Do initial estimation procedure to more accurately weight reads with multiple genome mappings
type: basic:boolean
description: Run an initial estimation procedure that weights reads mapping to multiple locations more accurately.
default: False

cxb

label: Abundances (.cxb)
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

Cuffquant results¶

data:cufflinks:cuffquantupload-cxb (basic:file src, basic:string source, basic:string species, basic:string build, basic:string feature_type)[Source: v1.3.2]

Upload Cuffquant results file (.cxb)

src

label: Cuffquant file
type: basic:file
description: Upload Cuffquant results file. Supported extention: *.cxb
required: True
validate_regex: \.(cxb)$

source

label

Gene ID database

type

basic:string

choices

AFFY: AFFY
DICTYBASE: DICTYBASE
ENSEMBL: ENSEMBL
NCBI: NCBI
UCSC: UCSC

species

label

Species

type

basic:string

description

Species latin name.

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Build
type: basic:string

feature_type

label

Feature type

type

basic:string

default

gene

choices

gene: gene
transcript: transcript
exon: exon

cxb

label: Cuffquant results
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

feature_type

label: Feature type
type: basic:string

Custom master file¶

data:masterfile:ampliconupload-master-file (basic:file src, basic:string panel_name)[Source: v1.2.1]

This should be a tab delimited file (*.txt). Please check the [example](http://genial.is/amplicon-masterfile) file for details.

src

label: Master file
type: basic:file
validate_regex: \.txt(|\.gz|\.bz2|\.tgz|\.tar\.gz|\.tar\.bz2|\.zip|\.rar|\.7z)$

panel_name

label: Panel name
type: basic:string

master_file

label: Master file
type: basic:file

bedfile

label: BED file (merged targets)
type: basic:file

nomergebed

label: BED file (nonmerged targets)
type: basic:file

olapfreebed

label: BED file (overlap-free targets)
type: basic:file

primers

label: Primers
type: basic:file

panel_name

label: Panel name
type: basic:string

Cut & Run¶

data:workflow:cutnrunworkflow-cutnrun (data:reads:fastq:paired reads, basic:integer quality, basic:integer nextseq, basic:string phred, basic:integer min_length, basic:integer max_n, basic:boolean retain_unpaired, basic:integer unpaired_len_1, basic:integer unpaired_len_2, basic:integer clip_r1, basic:integer clip_r2, basic:integer three_prime_r1, basic:integer three_prime_r2, list:basic:string adapter, list:basic:string adapter_2, data:seq:nucleotide adapter_file_1, data:seq:nucleotide adapter_file_2, basic:string universal_adapter, basic:integer stringency, basic:decimal error_rate, basic:integer trim_5, basic:integer trim_3, data:index:bowtie2 genome, basic:string mode, basic:string speed, basic:boolean discordantly, basic:boolean rep_se, basic:integer minins, basic:integer maxins, basic:boolean no_overlap, basic:boolean dovetail, basic:boolean no_unal, data:index:bowtie2 genome, basic:string mode, basic:string speed, basic:boolean discordantly, basic:boolean rep_se, basic:integer minins, basic:integer maxins, basic:boolean no_overlap, basic:boolean dovetail, basic:boolean no_unal, basic:string format, basic:decimal pvalue, basic:string duplicates, basic:boolean bedgraph, basic:integer min_frag_length, basic:integer max_frag_length, basic:decimal scale, basic:integer bw_binsize, basic:integer bw_timeout)[Source: v1.2.1]

Analysis of samples processed for high resolution mapping of DNA binding sites using targeted nuclease strategy. The process is named CUT&RUN which stands for Cleavage Under Target and Release Using Nuclease. Workflow includes steps of trimming the reads with trimgalore, aligning them using bowtie2 to target species genome as well as a spike-in genome. Aligned reads are processed to produce bigwig files to be viewed in a genome browser. Peaks are called using MACS2. Fragmenting of reads is performed using alignmentSieve from deeptools package.

reads

label: Input reads
type: data:reads:fastq:paired

options_trimming.quality_trim.quality

label: Quality cutoff
type: basic:integer
description: Trim low-quality ends from reads based on Phred score.
required: False

options_trimming.quality_trim.nextseq

label: NextSeq/NovaSeq trim cutoff
type: basic:integer
description: NextSeq/NovaSeq-specific quality trimming. Trims also dark cycles appearing as high-quality G bases. This will set a specific quality cutoff, but qualities of G bases are ignored. This can not be used with Quality cutoff and will override it.
required: False

options_trimming.quality_trim.phred

label

Phred score encoding

type

basic:string

description

Use either ASCII+33 quality scores as Phred scores (Sanger/Illumina 1 .9+ encoding) or ASCII+64 quality scores (Illumina 1.5 encoding) for quality trimming.

default

--phred33

choices

ASCII+33: --phred33
ASCII+64: --phred64

options_trimming.quality_trim.min_length

label: Minimum length after trimming
type: basic:integer
description: Discard reads that became shorter than selected length because of either quality or adapter trimming. Both reads of a read-pair need to be longer than specified length to be printed out to validated paired-end files. If only one read became too short there is the possibility of keeping such unpaired single-end reads with Retain unpaired. A value of 0 disables filtering based on length.
default: 20

options_trimming.quality_trim.max_n

label: Maximum number of Ns
type: basic:integer
description: Read exceeding this limit will result in the entire pair being removed from the trimmed output files.
required: False

options_trimming.quality_trim.retain_unpaired

label: Retain unpaired reads after trimming
type: basic:boolean
description: If only one of the two paired-end reads “became too short, the longer read will be written.
default: False

options_trimming.quality_trim.unpaired_len_1

label: Unpaired read length cutoff of mate 1
type: basic:integer
hidden: !quality_trim.retain_unpaired
default: 35

options_trimming.quality_trim.unpaired_len_2

label: Unpaired read length cutoff for mate 2
type: basic:integer
hidden: !quality_trim.retain_unpaired
default: 35

options_trimming.quality_trim.clip_r1

label: Trim bases from 5’ end of read 1
type: basic:integer
description: This may be useful if the qualities were very poor, or if there is some sort of unwanted bias at the 5’ end.
required: False

options_trimming.quality_trim.clip_r2

label: Trim bases from 5’ end of read 2
type: basic:integer
description: This may be useful if the qualities were very poor, or if there is some sort of unwanted bias at the 5’ end. For paired-end bisulfite sequencing, it is recommended to remove the first few bp because the end-repair reaction may introduce a bias towards low methylation.
required: False

options_trimming.quality_trim.three_prime_r1

label: Trim bases from 3’ end of read 1
type: basic:integer
description: Remove bases from the 3’ end of read 1 after adapter/quality trimming has been performed. This may remove some unwanted bias from the 3’ end that is not directly related to adapter sequence or basecall quality.
required: False

options_trimming.quality_trim.three_prime_r2

label: Trim bases from 3’ end of read 2
type: basic:integer
description: Remove bases from the 3’ end of read 2 after adapter/quality trimming has been performed. This may remove some unwanted bias from the 3’ end that is not directly related to adapter sequence or basecall quality.
required: False

options_trimming.adapter_trim.adapter

label: Read 1 adapter sequence
type: list:basic:string
description: Adapter sequences to be trimmed. Also see universal adapters field for predefined adapters. This is mutually exclusive with read 1 adapters file and universal adapters.
required: False

options_trimming.adapter_trim.adapter_2

label: Read 2 adapter sequence
type: list:basic:string
description: Optional adapter sequence to be trimmed off read 2 of paired-end files. This is mutually exclusive with read 2 adapters file and universal adapters.
required: False

options_trimming.adapter_trim.adapter_file_1

label: Read 1 adapters file
type: data:seq:nucleotide
description: This is mutually exclusive with read 1 adapters and universal adapters.
required: False

options_trimming.adapter_trim.adapter_file_2

label: Read 2 adapters file
type: data:seq:nucleotide
description: This is mutually exclusive with read 2 adapters and universal adapters.
required: False

options_trimming.adapter_trim.universal_adapter

label

Universal adapters

type

basic:string

description

Instead of default detection use specific adapters. Use 13bp of the Illumina universal adapter, 12bp of the Nextera adapter or 12bp of the Illumina Small RNA 3’ Adapter. Selecting to trim smallRNA adapters will also lower the length value to 18bp. If the smallRNA libraries are paired-end then read 2 adapter will be set to the Illumina small RNA 5’ adapter automatically (GATCGTCGGACT) unless defined explicitly. This is mutually exclusive with manually defined adapters and adapter files.

required

False

choices

Illumina: --illumina
Nextera: --nextera
Illumina small RNA: --small_rna

options_trimming.adapter_trim.stringency

label: Overlap with adapter sequence required to trim
type: basic:integer
description: Defaults to a very stringent setting of 1, i.e. even a single base pair of overlapping sequence will be trimmed of the 3’ end of any read.
default: 1

options_trimming.adapter_trim.error_rate

label: Maximum allowed error rate
type: basic:decimal
description: Number of errors divided by the length of the matching region. Default value of 0.1.
default: 0.1

options_trimming.hard_trim.trim_5

label: Hard trim sequence from 3’ end
type: basic:integer
description: Instead of performing adapter-/quality trimming, this option will simply hard-trim sequences to bp from the 3’ end. This is incompatible with other hard trimming options.
required: False

options_trimming.hard_trim.trim_3

label: Hard trim sequences from 5’ end
type: basic:integer
description: Instead of performing adapter-/quality trimming, this option will simply hard-trim sequences to bp from the 5’ end. This is incompatible with other hard trimming options.
required: False

options_aln_species.genome

label: Species genome
type: data:index:bowtie2

options_aln_species.mode

label

Alignment mode

type

basic:string

description

End to end: Bowtie 2 requires that the entire read align from one end to the other, without any trimming (or “soft clipping”) of characters from either end. Local: Bowtie 2 does not require that the entire read align from one end to the other. Rather, some characters may be omitted (“soft clipped”) from the ends in order to achieve the greatest possible alignment score.

default

--local

choices

end to end mode: --end-to-end
local: --local

options_aln_species.speed

label

Speed vs. Sensitivity

type

basic:string

description

A quick setting for aligning fast or accurately. This option is a shortcut for parameters as follows: For –end-to-end: –very-fast -D 5 -R 1 -N 0 -L 22 -i S,0,2.50 –fast -D 10 -R 2 -N 0 -L 22 -i S,0,2.50 –sensitive -D 15 -R 2 -N 0 -L 22 -i S,1,1.15 (default) –very-sensitive -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 For –local: –very-fast-local -D 5 -R 1 -N 0 -L 25 -i S,1,2.00 –fast-local -D 10 -R 2 -N 0 -L 22 -i S,1,1.75 –sensitive-local -D 15 -R 2 -N 0 -L 20 -i S,1,0.75 (default) –very-sensitive-local -D 20 -R 3 -N 0 -L 20 -i S,1,0.50

default

--very-sensitive

choices

Very fast: --very-fast
Fast: --fast
Sensitive: --sensitive
Very sensitive: --very-sensitive

options_aln_species.discordantly

label: Report discordantly matched read
type: basic:boolean
description: If both mates have unique alignments, but the alignments do not match paired-end expectations (orientation and relative distance) then alignment will be reported. Useful for detecting structural variations.
default: True

options_aln_species.rep_se

label: Report single ended
type: basic:boolean
description: If paired alignment can not be found Bowtie2 tries to find alignments for the individual mates. Default is true (–no-mixed).
default: True

options_aln_species.minins

label: Minimal distance
type: basic:integer
description: The minimum fragment length (–minins) for valid paired-end alignments. Value 0 imposes no minimum.
default: 10

options_aln_species.maxins

label: Maximal distance
type: basic:integer
description: The maximum fragment length (–maxins) for valid paired-end alignments.
default: 700

options_aln_species.no_overlap

label: Not concordant when mates overlap
type: basic:boolean
description: When true, it is considered not concordant when mates overlap at all. Defaul is true (–no-overlap).
default: False

options_aln_species.dovetail

label: Dovetail
type: basic:boolean
description: If the mates “dovetail”, that is if one mate alignment extends past the beginning of the other such that the wrong mate begins upstream, consider that to be concordant. Default: mates cannot dovetail in a concordant alignment. If true, parameter –dovetail is turned on.
default: False

options_aln_species.no_unal

label: Suppress SAM records for unaligned reads
type: basic:boolean
description: When true, suppress SAM records for unaligned reads. Default is true (–no-unal).
default: True

options_aln_spikein.genome

label: Spike-in genome
type: data:index:bowtie2

options_aln_spikein.mode

label

Alignment mode

type

basic:string

description

End to end: Bowtie 2 requires that the entire read align from one end to the other, without any trimming (or “soft clipping”) of characters from either end. Local: Bowtie 2 does not require that the entire read align from one end to the other. Rather, some characters may be omitted (“soft clipped”) from the ends in order to achieve the greatest possible alignment score.

default

--local

choices

end to end mode: --end-to-end
local: --local

options_aln_spikein.speed

label

Speed vs. Sensitivity

type

basic:string

description

A quick setting for aligning fast or accurately. This option is a shortcut for parameters as follows: For –end-to-end: –very-fast -D 5 -R 1 -N 0 -L 22 -i S,0,2.50 –fast -D 10 -R 2 -N 0 -L 22 -i S,0,2.50 –sensitive -D 15 -R 2 -N 0 -L 22 -i S,1,1.15 (default) –very-sensitive -D 20 -R 3 -N 0 -L 20 -i S,1,0.50 For –local: –very-fast-local -D 5 -R 1 -N 0 -L 25 -i S,1,2.00 –fast-local -D 10 -R 2 -N 0 -L 22 -i S,1,1.75 –sensitive-local -D 15 -R 2 -N 0 -L 20 -i S,1,0.75 (default) –very-sensitive-local -D 20 -R 3 -N 0 -L 20 -i S,1,0.50

default

--very-sensitive

choices

Very fast: --very-fast
Fast: --fast
Sensitive: --sensitive
Very sensitive: --very-sensitive

options_aln_spikein.discordantly

label: Report discordantly matched read
type: basic:boolean
description: If both mates have unique alignments, but the alignments do not match paired-end expectations (orientation and relative distance) then alignment will be reported. Useful for detecting structural variations.
default: True

options_aln_spikein.rep_se

label: Report single ended
type: basic:boolean
description: If paired alignment can not be found Bowtie2 tries to find alignments for the individual mates. Default is true (–no-mixed).
default: True

options_aln_spikein.minins

label: Minimal distance
type: basic:integer
description: The minimum fragment length (–minins) for valid paired-end alignments. Value 0 imposes no minimum.
default: 10

options_aln_spikein.maxins

label: Maximal distance
type: basic:integer
description: The maximum fragment length (–maxins) for valid paired-end alignments.
default: 700

options_aln_spikein.no_overlap

label: Not concordant when mates overlap
type: basic:boolean
description: When true, it is considered not concordant when mates overlap at all. Defaul is true (–no-overlap).
default: True

options_aln_spikein.dovetail

label: Dovetail
type: basic:boolean
description: If the mates “dovetail”, that is if one mate alignment extends past the beginning of the other such that the wrong mate begins upstream, consider that to be concordant. Default: mates cannot dovetail in a concordant alignment. If true, parameter –dovetail is turned on.
default: False

options_aln_spikein.no_unal

label: Suppress SAM records for unaligned reads
type: basic:boolean
description: When true, suppress SAM records for unaligned reads. Default is true (–no-unal).
default: True

options_pc.format

label

Format of tag file

type

basic:string

description

This specifies the format of input files. For paired-end data the format dicates how MACS2 will treat mates. If the selected format is BAM, MACS2 will only keep the left mate (5’ end) tag. However, when format BAMPE is selected, MACS2 will use actual insert sizes of pairs of reads to build fragment pileup, instead of building bimodal distribution plus and minus strand reads to predict fragment size.

required

False

default

BAMPE

choices

BAM: BAM
BAMPE: BAMPE

options_pc.pvalue

label: P-value cutoff
type: basic:decimal
description: The p-value cutoff.
required: False
default: 0.001

options_pc.duplicates

label

Number of duplicates

type

basic:string

description

It controls the MACS behavior towards duplicate tags at the exact same location – the same coordination and the same strand. The ‘auto’ option makes MACS calculate the maximum tags at the exact same location based on binomal distribution using 1e-5 as pvalue cutoff and the ‘all’ option keeps all the tags. If an integer is given, at most this number of tags will be kept at the same location. The default is to keep one tag at the same location.

default

all

choices

1: 1
auto: auto
all: all

options_pc.bedgraph

label: Save fragment pileup and control lambda
type: basic:boolean
description: If this flag is on, MACS will store the fragment pileup, control lambda, -log10(pvalue) and -log10(qvalue) scores in bedGraph files. The bedGraph files will be stored in current directory named NAME+’_treat_pileup.bdg’ for treatment data, NAME+’_control_lambda.bdg’ for local lambda values from control, NAME+’_treat_pvalue.bdg’ for Poisson pvalue scores (in -log10(pvalue) form), and NAME+’_treat_qvalue.bdg’ for q-value scores from Benjamini-Hochberg-Yekutieli procedure.
default: True

options_sieve.min_frag_length

label: Minimum fragment length
type: basic:integer
description: The minimum fragment length needed for read/pair inclusion. This option is primarily useful in ATACseq experiments, for filtering mono- or di-nucleosome fragments. Default is 0.
default: 0

options_sieve.max_frag_length

label: Maximum fragment length
type: basic:integer
description: The maximum fragment length needed for read/pair inclusion. A value of 0 indicates no limit. Default is 0.
default: 0

options_scale.scale

label: Scale factor
type: basic:decimal
description: Magnitude of the scale factor. The scaling factor is calculated by dividing the scale with the number of features in BEDPE (scale/(number of features)).
default: 10000

options_misc.bw_binsize

label: BigWig bin size
type: basic:integer
description: Size of the bins, in bases, for the output of the bigwig/bedgraph file. Default is 50.
default: 50

options_misc.bw_timeout

label: BigWig timeout
type: basic:integer
description: Number of seconds before calculation of BigWig file is aborted. Default is 3600 seconds (1 hour).
default: 3600

Cutadapt (3’ mRNA-seq, single-end)¶

data:reads:fastq:single:cutadapt:cutadapt-3prime-single (data:reads:fastq:single reads, basic:integer nextseq_trim, basic:integer quality_cutoff, basic:integer min_len, basic:integer min_overlap, basic:integer times)[Source: v1.2.1]

Process 3’ mRNA-seq datasets using Cutadapt tool.

reads

label: Select sample(s)
type: data:reads:fastq:single
required: True
hidden: False

options.nextseq_trim

label: NextSeq/NovaSeq trim
type: basic:integer
description: NextSeq/NovaSeq-specific quality trimming. Trims also dark cycles appearing as high-quality G bases. This option is mutually exclusive with the use of standard quality-cutoff trimming and is suitable for the use with data generated by the recent Illumina machines that utilize two-color chemistry to encode the four bases.
required: True
hidden: False
default: 10

options.quality_cutoff

label: Quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. The use of this option will override the use of NextSeq/NovaSeq trim option.
required: False
hidden: False

options.min_len

label: Discard reads shorter than specified minimum length.
type: basic:integer
required: True
hidden: False
default: 20

options.min_overlap

label: Mimimum overlap
type: basic:integer
description: Minimum overlap between adapter and read for an adapter to be found.
required: True
hidden: False
default: 20

options.times

label: Remove up to a specified number of adapters from each read.
type: basic:integer
required: True
hidden: False
default: 2

fastq

label: Reads file.
type: list:basic:file
required: True
hidden: False

report

label: Cutadapt report
type: basic:file
required: True
hidden: False

fastqc_url

label: Quality control with FastQC.
type: list:basic:file:html
required: True
hidden: False

fastqc_archive

label: Download FastQC archive.
type: list:basic:file
required: True
hidden: False

Cutadapt (Corall RNA-Seq, paired-end)¶

data:reads:fastq:paired:cutadapt:cutadapt-corall-paired (data:reads:fastq:paired reads, basic:integer nextseq_trim, basic:integer quality_cutoff, basic:integer min_len, basic:integer min_overlap)[Source: v1.1.2]

Pre-process reads obtained using CORALL Total RNA-Seq Library Prep Kit. Trim UMI-tags from input reads and use Cutadapt to remove adapters and run QC filtering steps.

reads

label: Select sample(s)
type: data:reads:fastq:paired
required: True
hidden: False

options.nextseq_trim

label: NextSeq/NovaSeq trim
type: basic:integer
description: NextSeq/NovaSeq-specific quality trimming. Trims also dark cycles appearing as high-quality G bases. This option is mutually exclusive with the use of standard quality-cutoff trimming and is suitable for the use with data generated by the recent Illumina machines that utilize two-color chemistry to encode the four bases.
required: True
hidden: False
default: 10

options.quality_cutoff

label: Quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. The use of this option will override the use of NextSeq/NovaSeq trim option.
required: False
hidden: False

options.min_len

label: Minimum read length
type: basic:integer
required: True
hidden: False
default: 20

options.min_overlap

label: Mimimum overlap
type: basic:integer
description: Minimum overlap between adapter and read for an adapter to be found.
required: True
hidden: False
default: 20

fastq

label: Remaining mate1 reads
type: list:basic:file
required: True
hidden: False

fastq2

label: Remaining mate2 reads
type: list:basic:file
required: True
hidden: False

report

label: Cutadapt report
type: basic:file
required: True
hidden: False

fastqc_url

label: Mate1 quality control with FastQC
type: list:basic:file:html
required: True
hidden: False

fastqc_url2

label: Mate2 quality control with FastQC
type: list:basic:file:html
required: True
hidden: False

fastqc_archive

label: Download mate1 FastQC archive
type: list:basic:file
required: True
hidden: False

fastqc_archive2

label: Download mate2 FastQC archive
type: list:basic:file
required: True
hidden: False

Cutadapt (Corall RNA-Seq, single-end)¶

data:reads:fastq:single:cutadapt:cutadapt-corall-single (data:reads:fastq:single reads, basic:integer nextseq_trim, basic:integer quality_cutoff, basic:integer min_len, basic:integer min_overlap)[Source: v1.2.1]

Pre-process reads obtained using CORALL Total RNA-Seq Library Prep Kit. Trim UMI-tags from input reads and use Cutadapt to remove adapters and run QC filtering steps.

reads

label: Select sample(s)
type: data:reads:fastq:single
required: True
hidden: False

options.nextseq_trim

label: NextSeq/NovaSeq trim
type: basic:integer
description: NextSeq/NovaSeq-specific quality trimming. Trims also dark cycles appearing as high-quality G bases. This option is mutually exclusive with the use of standard quality-cutoff trimming and is suitable for the use with data generated by the recent Illumina machines that utilize two-color chemistry to encode the four bases.
required: True
hidden: False
default: 10

options.quality_cutoff

label: Quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. The use of this option will override the use of NextSeq/NovaSeq trim option.
required: False
hidden: False

options.min_len

label: Minimum read length
type: basic:integer
required: True
hidden: False
default: 20

options.min_overlap

label: Mimimum overlap
type: basic:integer
description: Minimum overlap between adapter and read for an adapter to be found.
required: True
hidden: False
default: 20

fastq

label: Reads file
type: list:basic:file
required: True
hidden: False

report

label: Cutadapt report
type: basic:file
required: True
hidden: False

fastqc_url

label: Quality control with FastQC
type: list:basic:file:html
required: True
hidden: False

fastqc_archive

label: Download FastQC archive
type: list:basic:file
required: True
hidden: False

Cutadapt (Diagenode CATS, paired-end)¶

data:reads:fastq:paired:cutadaptcutadapt-custom-paired (data:reads:fastq:paired reads)[Source: v1.3.1]

Cutadapt process configured to be used with the Diagenode CATS kits.

reads

label: NGS reads
type: data:reads:fastq:paired

fastq

label: Reads file (forward)
type: list:basic:file

fastq2

label: Reads file (reverse)
type: list:basic:file

report

label: Cutadapt report
type: basic:file

fastqc_url

label: Quality control with FastQC (forward)
type: list:basic:file:html

fastqc_url2

label: Quality control with FastQC (reverse)
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive (forward)
type: list:basic:file

fastqc_archive2

label: Download FastQC archive (reverse)
type: list:basic:file

Cutadapt (Diagenode CATS, single-end)¶

data:reads:fastq:single:cutadaptcutadapt-custom-single (data:reads:fastq:single reads)[Source: v1.3.1]

Cutadapt process configured to be used with the Diagenode CATS kits.

reads

label: NGS reads
type: data:reads:fastq:single

fastq

label: Reads file
type: list:basic:file

report

label: Cutadapt report
type: basic:file

fastqc_url

label: Quality control with FastQC
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive
type: list:basic:file

Cutadapt (paired-end)¶

data:reads:fastq:paired:cutadaptcutadapt-paired (data:reads:fastq:paired reads, data:seq:nucleotide mate1_5prime_file, data:seq:nucleotide mate1_3prime_file, data:seq:nucleotide mate2_5prime_file, data:seq:nucleotide mate2_3prime_file, list:basic:string mate1_5prime_seq, list:basic:string mate1_3prime_seq, list:basic:string mate2_5prime_seq, list:basic:string mate2_3prime_seq, basic:integer times, basic:decimal error_rate, basic:integer min_overlap, basic:boolean match_read_wildcards, basic:integer nextseq_trim, basic:integer leading, basic:integer trailing, basic:integer crop, basic:integer headcrop, basic:integer minlen, basic:integer max_n, basic:string pair_filter)[Source: v2.4.1]

Cutadapt finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. More information about Cutadapt can be found [here](http://cutadapt.readthedocs.io/en/stable/).

reads

label: Select sample(s)
type: data:reads:fastq:paired

adapters.mate1_5prime_file

label: 5 prime adapter file for Mate 1
type: data:seq:nucleotide
required: False

adapters.mate1_3prime_file

label: 3 prime adapter file for Mate 1
type: data:seq:nucleotide
required: False

adapters.mate2_5prime_file

label: 5 prime adapter file for Mate 2
type: data:seq:nucleotide
required: False

adapters.mate2_3prime_file

label: 3 prime adapter file for Mate 2
type: data:seq:nucleotide
required: False

adapters.mate1_5prime_seq

label: 5 prime adapter sequence for Mate 1
type: list:basic:string
required: False

adapters.mate1_3prime_seq

label: 3 prime adapter sequence for Mate 1
type: list:basic:string
required: False

adapters.mate2_5prime_seq

label: 5 prime adapter sequence for Mate 2
type: list:basic:string
required: False

adapters.mate2_3prime_seq

label: 3 prime adapter sequence for Mate 2
type: list:basic:string
required: False

adapters.times

label: Times
type: basic:integer
description: Remove up to COUNT adapters from each read.
default: 1

adapters.error_rate

label: Error rate
type: basic:decimal
description: Maximum allowed error rate (no. of errors divided by the length of the matching region).
default: 0.1

adapters.min_overlap

label: Minimal overlap
type: basic:integer
description: Minimum overlap for an adapter match.
default: 3

adapters.match_read_wildcards

label: Match read wildcards
type: basic:boolean
description: Interpret IUPAC wildcards in reads.
default: False

modify_reads.nextseq_trim

label: NextSeq-specific quality trimming
type: basic:integer
description: NextSeq-specific quality trimming (each read). Trims also dark cycles appearing as high-quality G bases. This option is mutually exclusive with the use of regular (-g) quality trimming.
required: False

modify_reads.leading

label: Quality on 5 prime
type: basic:integer
description: Remove low quality bases from 5 prime. Specifies the minimum quality required to keep a base.
required: False

modify_reads.trailing

label: Quality on 3 prime
type: basic:integer
description: Remove low quality bases from the 3 prime. Specifies the minimum quality required to keep a base.
required: False

modify_reads.crop

label: Crop
type: basic:integer
description: Cut the specified number of bases from the end of the reads.
required: False

modify_reads.headcrop

label: Headcrop
type: basic:integer
description: Cut the specified number of bases from the start of the reads.
required: False

filtering.minlen

label: Min length
type: basic:integer
description: Drop the read if it is below a specified.
required: False

filtering.max_n

label: Max numebr of N-s
type: basic:integer
description: Discard reads having more ‘N’ bases than specified.
required: False

filtering.pair_filter

label

Which of the reads have to match the filtering criterion

type

basic:string

description

Which of the reads in a paired-end read have to match the filtering criterion in order for the pair to be filtered.

default

any

choices

Any of the reads in a paired-end read have to match the filtering criterion: any
Both of the reads in a paired-end read have to match the filtering criterion: both

fastq

label: Reads file (forward)
type: list:basic:file

fastq2

label: Reads file (reverse)
type: list:basic:file

report

label: Cutadapt report
type: basic:file

fastqc_url

label: Quality control with FastQC (forward)
type: list:basic:file:html

fastqc_url2

label: Quality control with FastQC (reverse)
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive (forward)
type: list:basic:file

fastqc_archive2

label: Download FastQC archive (reverse)
type: list:basic:file

Cutadapt (single-end)¶

data:reads:fastq:single:cutadaptcutadapt-single (data:reads:fastq:single reads, data:seq:nucleotide up_primers_file, data:seq:nucleotide down_primers_file, list:basic:string up_primers_seq, list:basic:string down_primers_seq, basic:integer polya_tail, basic:integer min_overlap, basic:integer nextseq_trim, basic:integer leading, basic:integer trailing, basic:integer crop, basic:integer headcrop, basic:integer minlen, basic:integer max_n, basic:boolean match_read_wildcards, basic:integer times, basic:decimal error_rate)[Source: v2.2.1]

Cutadapt finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. More information about Cutadapt can be found [here](http://cutadapt.readthedocs.io/en/stable/).

reads

label: Select sample(s)
type: data:reads:fastq:single

adapters.up_primers_file

label: 5 prime adapter file
type: data:seq:nucleotide
required: False

adapters.down_primers_file

label: 3 prime adapter file
type: data:seq:nucleotide
required: False

adapters.up_primers_seq

label: 5 prime adapter sequence
type: list:basic:string
required: False

adapters.down_primers_seq

label: 3 prime adapter sequence
type: list:basic:string
required: False

adapters.polya_tail

label: Poly-A tail
type: basic:integer
description: Length of poly-A tail, example - AAAN -> 3, AAAAAN -> 5
required: False

adapters.min_overlap

label: Minimal overlap
type: basic:integer
description: Minimum overlap for an adapter match
default: 3

modify_reads.nextseq_trim

label: NextSeq-specific quality trimming
type: basic:integer
description: NextSeq-specific quality trimming (each read). Trims also dark cycles appearing as high-quality G bases. This option is mutually exclusive with the use of regular (-g) quality trimming.
required: False

modify_reads.leading

label: Quality on 5 prime
type: basic:integer
description: Remove low quality bases from 5 prime. Specifies the minimum quality required to keep a base. This option is mutually exclusive with the use of NextSeq-specific quality trimming.
required: False

modify_reads.trailing

label: Quality on 3 prime
type: basic:integer
description: Remove low quality bases from the 3 prime. Specifies the minimum quality required to keep a base. This option is mutually exclusive with the use of NextSeq-specific quality trimming.
required: False

modify_reads.crop

label: Crop
type: basic:integer
description: Cut the read to a specified length by removing bases from the end
required: False

modify_reads.headcrop

label: Headcrop
type: basic:integer
description: Cut the specified number of bases from the start of the read
required: False

filtering.minlen

label: Min length
type: basic:integer
description: Drop the read if it is below a specified length
required: False

filtering.max_n

label: Max numebr of N-s
type: basic:integer
description: Discard reads having more ‘N’ bases than specified.
required: False

filtering.match_read_wildcards

label: Match read wildcards
type: basic:boolean
description: Interpret IUPAC wildcards in reads.
required: False
default: False

filtering.times

label: Times
type: basic:integer
description: Remove up to COUNT adapters from each read.
default: 1

filtering.error_rate

label: Error rate
type: basic:decimal
description: Maximum allowed error rate (no. of errors divided by the length of the matching region).
default: 0.1

fastq

label: Reads file
type: list:basic:file

report

label: Cutadapt report
type: basic:file

fastqc_url

label: Quality control with FastQC
type: list:basic:file:html

fastqc_archive

label: Download FastQC archive
type: list:basic:file

Cutadapt - STAR - FeatureCounts (3’ mRNA-Seq, single-end)¶

data:workflow:quant:featurecounts:singleworkflow-cutadapt-star-fc-quant-single (data:reads:fastq:single reads, data:index:star star_index, data:annotation annotation, data:index:star rrna_reference, data:index:star globin_reference, basic:boolean show_advanced, basic:integer quality_cutoff, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass)[Source: v2.0.1]

This 3’ mRNA-Seq pipeline is comprised of QC, preprocessing, alignment and quantification steps. Reads are preprocessed by __Cutadapt__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Preprocessed reads are aligned by __STAR__ aligner. For read-count quantification, the __FeatureCounts__ tool is used. QoRTs QC and Samtools idxstats tools are used to report alignment QC metrics. Additional QC steps operate on downsampled reads and include an alignment of input reads to the rRNA/globin reference sequences. The reported alignment rate is used to asses the rRNA/globin sequence depletion rate.

reads

label: Select sample(s)
type: data:reads:fastq:single

star_index

label: Genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation
description: Genome annotation file (GTF).

rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

cutadapt.quality_cutoff

label: Reads quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. The use of this option will override the use of NextSeq/NovaSeq-specific trim option.
required: False

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.seed

label: Seed
type: basic:integer
default: 11

downsampling.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads in range [0.0, 1.0] from the original input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

Cutadapt - STAR - FeatureCounts - basic QC (3’ mRNA-Seq, single-end)¶

data:workflow:quant:featurecounts:singleworkflow-cutadapt-star-fc-quant-wo-depletion-single (data:reads:fastq:single reads, data:index:star star_index, data:annotation annotation, basic:boolean show_advanced, basic:integer quality_cutoff)[Source: v2.0.1]

This 3’ mRNA-Seq pipeline is comprised of QC, preprocessing, alignment and quantification steps. Reads are preprocessed by __Cutadapt__ which removes adapters, trims reads for quality from the 3’-end, and discards reads that are too short after trimming. Preprocessed reads are aligned by __STAR__ aligner. For read-count quantification, the __FeatureCounts__ tool is used. QoRTs QC and Samtools idxstats tools are used to report alignment QC metrics.

reads

label: Select sample(s)
type: data:reads:fastq:single

star_index

label: Genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation
description: Genome annotation file (GTF).

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

cutadapt.quality_cutoff

label: Reads quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. The use of this option will override the use of NextSeq/NovaSeq-specific trim option.
required: False

Cutadapt - STAR - HTSeq-count (paired-end)¶

data:workflow:rnaseq:htseqworkflow-custom-cutadapt-star-htseq-paired (data:reads:fastq:paired reads, data:index:star genome, data:annotation:gtf gff, basic:string stranded, basic:boolean advanced, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax, basic:string mode, basic:string feature_class, basic:string id_attribute, basic:boolean name_ordered)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by __cutadapt__ which finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5792058/). Finally, aligned reads are summarized to genes by __HTSeq-count__. Compared to featureCounts, HTSeq-count is not as computationally efficient. All three tools in this workflow support parallelization to accelerate the analysis.

reads

label: NGS reads
type: data:reads:fastq:paired

genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool

gff

label: Annotation (GFF)
type: data:annotation:gtf

stranded

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

default

no

choices

Strand non-specific: no
Strand-specific forward: yes
Strand-specific reverse: reverse

advanced

label: Advanced
type: basic:boolean
default: False

star.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
default: False

star.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

star.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: !star.detect_chimeric.chimeric
default: 20

star.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: False

star.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: !star.t_coordinates.quantmode
default: False

star.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: !star.t_coordinates.quantmode
default: False

star.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

star.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

star.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

star.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

star.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

star.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

star.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

star.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

star.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

htseq.mode

label

Mode

type

basic:string

description

Mode to handle reads overlapping more than one feature. Possible values for <mode> are union, intersection-strict and intersection-nonempty

default

union

choices

union: union
intersection-strict: intersection-strict
intersection-nonempty: intersection-nonempty

htseq.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GFF file) to be used. All other features will be ignored.
default: exon

htseq.id_attribute

label: ID attribute
type: basic:string
description: GFF attribute to be used as feature ID. Several GFF lines with the same feature ID will be considered as parts of the same feature. The feature ID is used to identity the counts in the output table.
default: gene_id

htseq.name_ordered

label: Use name-ordered BAM file for counting reads
type: basic:boolean
description: Use name-sorted BAM file for reads quantification. Improves compatibility with larger BAM files, but requires more computational time.
required: False
default: False

Cutadapt - STAR - HTSeq-count (single-end)¶

data:workflow:rnaseq:htseqworkflow-custom-cutadapt-star-htseq-single (data:reads:fastq:single reads, data:index:star genome, data:annotation:gtf gff, basic:string stranded, basic:boolean advanced, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax, basic:string mode, basic:string feature_class, basic:string id_attribute, basic:boolean name_ordered)[Source: v2.0.1]

This RNA-seq pipeline is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by __cutadapt__ which finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. Next, preprocessed reads are aligned by __STAR__ aligner. At the time of implementation, STAR is considered a state-of-the-art tool that consistently produces accurate results from diverse sets of reads, and performs well even with default settings. For more information see [this comparison of RNA-seq aligners](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5792058/). Finally, aligned reads are summarized to genes by __HTSeq-count__. Compared to featureCounts, HTSeq-count is not as computationally efficient. All three tools in this workflow support parallelization to accelerate the analysis.

reads

label: NGS reads
type: data:reads:fastq:single

genome

label: Indexed reference genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool

gff

label: Annotation (GFF)
type: data:annotation:gtf

stranded

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

default

no

choices

Strand non-specific: no
Strand-specific forward: yes
Strand-specific reverse: reverse

advanced

label: Advanced
type: basic:boolean
default: False

star.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
default: False

star.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

star.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: !star.detect_chimeric.chimeric
default: 20

star.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: False

star.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: !star.t_coordinates.quantmode
default: False

star.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: !star.t_coordinates.quantmode
default: False

star.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

star.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

star.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

star.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

star.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

star.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

star.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

star.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

star.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

htseq.mode

label

Mode

type

basic:string

description

Mode to handle reads overlapping more than one feature. Possible values for <mode> are union, intersection-strict and intersection-nonempty

default

union

choices

union: union
intersection-strict: intersection-strict
intersection-nonempty: intersection-nonempty

htseq.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GFF file) to be used. All other features will be ignored.
default: exon

htseq.id_attribute

label: ID attribute
type: basic:string
description: GFF attribute to be used as feature ID. Several GFF lines with the same feature ID will be considered as parts of the same feature. The feature ID is used to identity the counts in the output table.
default: gene_id

htseq.name_ordered

label: Use name-ordered BAM file for counting reads
type: basic:boolean
description: Use name-sorted BAM file for reads quantification. Improves compatibility with larger BAM files, but requires more computational time.
required: False
default: False

Cutadapt - STAR - RSEM (Diagenode CATS, paired-end)¶

data:workflow:rnaseq:rsemworkflow-custom-cutadapt-star-rsem-paired (data:reads:fastq:paired reads, data:index:star star_index, data:index:expression expression_index, basic:string stranded, basic:boolean advanced, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax)[Source: v2.0.1]

This RNA-seq pipeline is configured to be used with the Diagenode CATS RNA-seq kits. It is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by cutadapt which finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. Next, preprocessed reads are aligned by STAR aligner. Finally, RSEM estimates gene and isoform expression levels from the aligned reads.

reads

label: NGS reads
type: data:reads:fastq:paired

star_index

label: STAR genome index
type: data:index:star

expression_index

label: Gene expression indices
type: data:index:expression

stranded

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

default

no

choices

Strand non-specific: no
Strand-specific forward: yes
Strand-specific reverse: reverse

advanced

label: Advanced
type: basic:boolean
default: False

star.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
default: False

star.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

star.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: !star.detect_chimeric.chimeric
default: 20

star.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: True

star.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: !star.t_coordinates.quantmode
default: False

star.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: !star.t_coordinates.quantmode
default: False

star.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

star.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

star.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

star.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

star.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

star.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

star.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

star.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

star.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

Cutadapt - STAR - RSEM (Diagenode CATS, single-end)¶

data:workflow:rnaseq:rsemworkflow-custom-cutadapt-star-rsem-single (data:reads:fastq:single reads, data:index:star star_index, data:index:expression expression_index, basic:string stranded, basic:boolean advanced, basic:boolean noncannonical, basic:boolean chimeric, basic:integer chimSegmentMin, basic:boolean quantmode, basic:boolean singleend, basic:boolean gene_counts, basic:string outFilterType, basic:integer outFilterMultimapNmax, basic:integer outFilterMismatchNmax, basic:decimal outFilterMismatchNoverLmax, basic:integer alignSJoverhangMin, basic:integer alignSJDBoverhangMin, basic:integer alignIntronMin, basic:integer alignIntronMax, basic:integer alignMatesGapMax)[Source: v2.0.1]

This RNA-seq pipeline is configured to be used with the Diagenode CATS RNA-seq kits. It is comprised of three steps, preprocessing, alignment, and quantification. First, reads are preprocessed by cutadapt which finds and removes adapter sequences, primers, poly-A tails and other types of unwanted sequence from high-throughput sequencing reads. Next, preprocessed reads are aligned by STAR aligner. Finally, RSEM estimates gene and isoform expression levels from the aligned reads.

reads

label: NGS reads
type: data:reads:fastq:single

star_index

label: STAR genome index
type: data:index:star

expression_index

label: Gene expression indices
type: data:index:expression

stranded

label

Assay type

type

basic:string

description

In strand non-specific assay a read is considered overlapping with a feature regardless of whether it is mapped to the same or the opposite strand as the feature. In strand-specific forward assay and single reads, the read has to be mapped to the same strand as the feature. For paired-end reads, the first read has to be on the same strand and the second read on the opposite strand. In strand-specific reverse assay these rules are reversed.

default

no

choices

Strand non-specific: no
Strand-specific forward: yes
Strand-specific reverse: reverse

advanced

label: Advanced
type: basic:boolean
default: False

star.noncannonical

label: Remove non-cannonical junctions (Cufflinks compatibility)
type: basic:boolean
description: It is recommended to remove the non-canonical junctions for Cufflinks runs using –outFilterIntronMotifs RemoveNoncanonical.
default: False

star.detect_chimeric.chimeric

label: Detect chimeric and circular alignments
type: basic:boolean
description: To switch on detection of chimeric (fusion) alignments (in addition to normal mapping), –chimSegmentMin should be set to a positive value. Each chimeric alignment consists of two “segments”. Each segment is non-chimeric on its own, but the segments are chimeric to each other (i.e. the segments belong to different chromosomes, or different strands, or are far from each other). Both segments may contain splice junctions, and one of the segments may contain portions of both mates. –chimSegmentMin parameter controls the minimum mapped length of the two segments that is allowed. For example, if you have 2x75 reads and used –chimSegmentMin 20, a chimeric alignment with 130b on one chromosome and 20b on the other will be output, while 135 + 15 won’t be.
default: False

star.detect_chimeric.chimSegmentMin

label: –chimSegmentMin
type: basic:integer
disabled: !star.detect_chimeric.chimeric
default: 20

star.t_coordinates.quantmode

label: Output in transcript coordinates
type: basic:boolean
description: With –quantMode TranscriptomeSAM option STAR will output alignments translated into transcript coordinates in the Aligned.toTranscriptome.out.bam file (in addition to alignments in genomic coordinates in Aligned.*.sam/bam files). These transcriptomic alignments can be used with various transcript quantification software that require reads to be mapped to transcriptome, such as RSEM or eXpress.
default: True

star.t_coordinates.singleend

label: Allow soft-clipping and indels
type: basic:boolean
description: By default, the output satisfies RSEM requirements: soft-clipping or indels are not allowed. Use –quantTranscriptomeBan Singleend to allow insertions, deletions ans soft-clips in the transcriptomic alignments, which can be used by some expression quantification software (e.g. eXpress).
disabled: !star.t_coordinates.quantmode
default: False

star.t_coordinates.gene_counts

label: Count reads
type: basic:boolean
description: With –quantMode GeneCounts option STAR will count number reads per gene while mapping. A read is counted if it overlaps (1nt or more) one and only one gene. Both ends of the paired-end read are checked for overlaps. The counts coincide with those produced by htseq-count with default parameters. ReadsPerGene.out.tab file with 4 columns which correspond to different strandedness options: column 1: gene ID; column 2: counts for unstranded RNA-seq; column 3: counts for the 1st read strand aligned with RNA (htseq-count option -s yes); column 4: counts for the 2nd read strand aligned with RNA (htseq-count option -s reverse).
disabled: !star.t_coordinates.quantmode
default: False

star.filtering.outFilterType

label

Type of filtering

type

basic:string

description

Normal: standard filtering using only current alignment; BySJout: keep only those reads that contain junctions that passed filtering into SJ.out.tab

default

Normal

choices

Normal: Normal
BySJout: BySJout

star.filtering.outFilterMultimapNmax

label: –outFilterMultimapNmax
type: basic:integer
description: Read alignments will be output only if the read maps fewer than this value, otherwise no alignments will be output (default: 10).
required: False

star.filtering.outFilterMismatchNmax

label: –outFilterMismatchNmax
type: basic:integer
description: Alignment will be output only if it has fewer mismatches than this value (default: 10).
required: False

star.filtering.outFilterMismatchNoverLmax

label: –outFilterMismatchNoverLmax
type: basic:decimal
description: Max number of mismatches per pair relative to read length: for 2x100b, max number of mismatches is 0.06*200=8 for the paired read.
required: False

star.alignment.alignSJoverhangMin

label: –alignSJoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for spliced alignments (default: 5).
required: False

star.alignment.alignSJDBoverhangMin

label: –alignSJDBoverhangMin
type: basic:integer
description: Minimum overhang (i.e. block size) for annotated (sjdb) spliced alignments (default: 3).
required: False

star.alignment.alignIntronMin

label: –alignIntronMin
type: basic:integer
description: Minimum intron size: genomic gap is considered intron if its length >= alignIntronMin, otherwise it is considered Deletion (default: 21).
required: False

star.alignment.alignIntronMax

label: –alignIntronMax
type: basic:integer
description: Maximum intron size, if 0, max intron size will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

star.alignment.alignMatesGapMax

label: –alignMatesGapMax
type: basic:integer
description: Maximum gap between two mates, if 0, max intron gap will be determined by (2pow(winBinNbits)*winAnchorDistNbins) (default: 0).
required: False

Cutadapt - STAR - StringTie (Corall, paired-end)¶

data:workflow:rnaseq:corallworkflow-corall-paired (data:reads:fastq:paired reads, data:index:star star_index, data:annotation annotation, data:index:star rrna_reference, data:index:star globin_reference, basic:boolean show_advanced, basic:integer quality_cutoff, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, basic:string feature_class, basic:string id_attribute)[Source: v3.0.1]

RNA-seq pipeline optimized for the Lexogen Corall Total RNA-Seq Library Prep Kit. UMI-sequences are extracted from the raw reads before the reads are trimmed and quality filtered using Cutadapt. Preprocessed reads are aligned by the STAR aligner and de-duplicated using UMI-tools. Gene abundance estimates are reported by the featureCounts tool. QC operates on downsampled reads and includes alignment of input reads to the rRNA/globin reference sequences. The reported alignment rate is used to asses the rRNA/globin sequence depletion rate. The analysis results and QC reports are summarized by the MultiQC.

reads

label: Select sample(s)
type: data:reads:fastq:paired

star_index

label: Genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation
description: Genome annotation file (GTF).

rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

cutadapt.quality_cutoff

label: Reads quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. Use this option when processing the data generated by older Illumina machines. The use of this option will override the NextSeq/NovaSeq-specific trimming procedure which is enabled by default and is recommended for Illumina machines that utilize 2-color chemistry to encode the four bases.
required: False

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.seed

label: Seed
type: basic:integer
default: 11

downsampling.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads in range [0.0, 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

quantification.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GTF/GFF3 file) to be used. All other features will be ignored.
default: exon

quantification.id_attribute

label

ID attribute

type

basic:string

description

GTF/GFF3 attribute to be used as feature ID. Several GTF/GFF3 lines with the same feature ID will be considered as parts of the same feature. The feature ID is used to identify the counts in the output table. In GTF files this is usually ‘gene_id’, in GFF3 files this is often ‘ID’, and ‘transcript_id’ is frequently a valid choice for both annotation formats.

default

gene_id

choices

gene_id: gene_id
transcript_id: transcript_id
ID: ID
geneid: geneid

Cutadapt - STAR - StringTie (Corall, single-end)¶

data:workflow:rnaseq:corallworkflow-corall-single (data:reads:fastq:single reads, data:index:star star_index, data:annotation annotation, data:index:star rrna_reference, data:index:star globin_reference, basic:boolean show_advanced, basic:integer quality_cutoff, basic:integer n_reads, basic:integer seed, basic:decimal fraction, basic:boolean two_pass, basic:string feature_class, basic:string id_attribute)[Source: v3.0.1]

RNA-seq pipeline optimized for the Lexogen Corall Total RNA-Seq Library Prep Kit. UMI-sequences are extracted from the raw reads before the reads are trimmed and quality filtered using Cutadapt. Preprocessed reads are aligned by the STAR aligner and de-duplicated using UMI-tools. Gene abundance estimates are reported by the featureCounts tool. QC operates on downsampled reads and includes alignment of input reads to the rRNA/globin reference sequences. The reported alignment rate is used to asses the rRNA/globin sequence depletion rate. The analysis results and QC reports are summarized by the MultiQC.

reads

label: Select sample(s)
type: data:reads:fastq:single

star_index

label: Genome
type: data:index:star
description: Genome index prepared by STAR aligner indexing tool.

annotation

label: Annotation
type: data:annotation
description: Genome annotation file (GTF).

rrna_reference

label: Indexed rRNA reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

globin_reference

label: Indexed Globin reference sequence
type: data:index:star
description: Reference sequence index prepared by STAR aligner indexing tool.

show_advanced

label: Show advanced parameters
type: basic:boolean
default: False

cutadapt.quality_cutoff

label: Reads quality cutoff
type: basic:integer
description: Trim low-quality bases from 3’ end of each read before adapter removal. Use this option when processing the data generated by older Illumina machines. The use of this option will override the NextSeq/NovaSeq-specific trimming procedure which is enabled by default and is recommended for Illumina machines that utilize 2-color chemistry to encode the four bases.
required: False

downsampling.n_reads

label: Number of reads
type: basic:integer
default: 1000000

downsampling.seed

label: Seed
type: basic:integer
default: 11

downsampling.fraction

label: Fraction
type: basic:decimal
description: Use the fraction of reads in range [0.0, 1.0] from the orignal input file instead of the absolute number of reads. If set, this will override the “Number of reads” input parameter.
required: False

downsampling.two_pass

label: 2-pass mode
type: basic:boolean
description: Enable two-pass mode when down-sampling. Two-pass mode is twice as slow but with much reduced memory.
default: False

quantification.feature_class

label: Feature class
type: basic:string
description: Feature class (3rd column in GTF/GFF3 file) to be used. All other features will be ignored.
default: exon

quantification.id_attribute

label

ID attribute

type

basic:string

description

GTF/GFF3 attribute to be used as feature ID. Several GTF/GFF3 lines with the same feature ID will be considered as parts of the same feature. The feature ID is used to identify the counts in the output table. In GTF files this is usually ‘gene_id’, in GFF3 files this is often ‘ID’, and ‘transcript_id’ is frequently a valid choice for both annotation formats.

default

gene_id

choices

gene_id: gene_id
transcript_id: transcript_id
ID: ID
geneid: geneid

DESeq2¶

data:differentialexpression:deseq2:differentialexpression-deseq2 (list:data:expression case, list:data:expression control, basic:boolean create_sets, basic:decimal logfc, basic:decimal fdr, basic:boolean beta_prior, basic:boolean count, basic:integer min_count_sum, basic:boolean cook, basic:decimal cooks_cutoff, basic:boolean independent, basic:decimal alpha)[Source: v3.2.2]

Run DESeq2 analysis. The DESeq2 package estimates variance-mean dependence in count data from high-throughput sequencing assays and tests for differential expression based on a model using the negative binomial distribution. See [here](https://www.bioconductor.org/packages/release/bioc/manuals/DESeq2/man/DESeq2.pdf) and [here](http://bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html) for more information.

case

label: Case
type: list:data:expression
description: Case samples (replicates)
required: True
hidden: False

control

label: Control
type: list:data:expression
description: Control samples (replicates)
required: True
hidden: False

create_sets

label: Create gene sets
type: basic:boolean
description: After calculating differential gene expressions create gene sets for up-regulated genes, down-regulated genes and all genes.
required: True
hidden: False
default: False

logfc

label: Log2 fold change threshold for gene sets
type: basic:decimal
description: Genes above Log2FC are considered as up-regulated and genes below -Log2FC as down-regulated.
required: True
hidden: !create_sets
default: 1.0

fdr

label: FDR threshold for gene sets
type: basic:decimal
required: True
hidden: !create_sets
default: 0.05

options.beta_prior

label: Beta prior
type: basic:boolean
description: Whether or not to put a zero-mean normal prior on the non-intercept coefficients.
required: True
hidden: False
default: False

filter_options.count

label: Filter genes based on expression count
type: basic:boolean
required: True
hidden: False
default: True

filter_options.min_count_sum

label: Minimum gene expression count summed over all samples
type: basic:integer
description: Filter genes in the expression matrix input. Remove genes where the expression count sum over all samples is below the threshold.
required: True
hidden: !filter_options.count
default: 10

filter_options.cook

label: Filter genes based on Cook’s distance
type: basic:boolean
required: True
hidden: False
default: False

filter_options.cooks_cutoff

label: Threshold on Cook’s distance
type: basic:decimal
description: If one or more samples have Cook’s distance larger than the threshold set here, the p-value for the row is set to NA. If left empty, the default threshold of 0.99 quantile of the F(p, m-p) distribution is used, where p is the number of coefficients being fitted and m is the number of samples. This test excludes Cook’s distance of samples belonging to experimental groups with only two samples.
required: False
hidden: !filter_options.cook

filter_options.independent

label: Apply independent gene filtering
type: basic:boolean
required: True
hidden: False
default: False

filter_options.alpha

label: Significance cut-off used for optimizing independent gene filtering
type: basic:decimal
description: The value should be set to adjusted p-value cut-off (FDR).
required: True
hidden: !filter_options.independent
default: 0.1

raw

label: Differential expression
type: basic:file
required: True
hidden: False

de_json

label: Results table (JSON)
type: basic:json
required: True
hidden: False

de_file

label: Results table (file)
type: basic:file
required: True
hidden: False

count_matrix

label: Count matrix
type: basic:file
required: True
hidden: False

source

label: Gene ID database
type: basic:string
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

feature_type

label: Feature type
type: basic:string
required: True
hidden: False

Deeptools bamCoverage¶

data:coverage:bigwig:scale-bigwig (data:alignment:bam alignment, data:bedpe bedpe, basic:decimal scale)[Source: v1.1.1]

Creates a scaled BigWig file.

alignment

label: Alignment BAM file
type: data:alignment:bam
required: True
hidden: False

bedpe

label: BEDPE Normalization factor
type: data:bedpe
description: The BEDPE file describes disjoint genome features, such as structural variations or paired-end sequence alignments. It is used to estimate the scale factor.
required: True
hidden: False

scale

label: Scale for the normalization factor
type: basic:decimal
description: Magnitude of the scale factor. The scaling factor is calculated by dividing the scale with the number of features in BEDPE (scale/(number of features)).
required: True
hidden: False
default: 10000

bigwig

label: bigwig file
type: basic:file
required: True
hidden: False

species

label: Species
type: basic:string
required: True
hidden: False

build

label: Build
type: basic:string
required: True
hidden: False

Detect library strandedness¶

data:strandednesslibrary-strandedness (data:reads:fastq reads, basic:integer read_number, data:index:salmon salmon_index)[Source: v0.4.1]

This process uses the Salmon transcript quantification tool to automatically infer the NGS library strandedness. For more details, please see the Salmon [documentation](https://salmon.readthedocs.io/en/latest/library_type.html)

reads

label: Sequencing reads
type: data:reads:fastq
description: Sequencing reads in .fastq format. Both single and paired-end libraries are supported

read_number

label: Number of input reads
type: basic:integer
description: Number of sequencing reads that are subsampled from each of the original .fastq files before library strand detection
default: 50000

salmon_index

label: Transcriptome index file
type: data:index:salmon
description: Transcriptome index file created using the Salmon indexing tool. cDNA (transcriptome) sequences used for index file creation must be derived from the same species as the input sequencing reads to obtain the reliable analysis results

strandedness

label: Library strandedness type
type: basic:string
description: The predicted library strandedness type. The codes U and IU indicate ‘strand non-specific’ library for single or paired-end reads, respectively. Codes SF and ISF correspond to the ‘strand-specific forward’ library, for the single or paired-end reads, respectively. For ‘strand-specific reverse’ library, the corresponding codes are SR and ISR. For more details, please see the Salmon [documentation](https://salmon.readthedocs.io/en/latest/library_type.html)

fragment_ratio

label: Compatible fragment ratio
type: basic:decimal
description: The ratio of fragments that support the predicted library strandedness type

log

label: Log file
type: basic:file
description: Analysis log file.

Dictyostelium expressions¶

data:expression:polyaexpression-dicty (data:alignment:bam alignment, data:annotation:gff3 gff, data:mappability:bcm mappable)[Source: v1.4.1]

Dictyostelium-specific pipeline. Developed by Bioinformatics Laboratory, Faculty of Computer and Information Science, University of Ljubljana, Slovenia and Shaulsky Lab, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, USA.

alignment

label: Aligned sequence
type: data:alignment:bam

gff

label: Features (GFF3)
type: data:annotation:gff3

mappable

label: Mappability
type: data:mappability:bcm

exp

label: Expression RPKUM (polyA)
type: basic:file
description: mRNA reads scaled by uniquely mappable part of exons.

rpkmpolya

label: Expression RPKM (polyA)
type: basic:file
description: mRNA reads scaled by exon length.

rc

label: Read counts (polyA)
type: basic:file
description: mRNA reads uniquely mapped to gene exons.

rpkum

label: Expression RPKUM
type: basic:file
description: Reads scaled by uniquely mappable part of exons.

rpkm

label: Expression RPKM
type: basic:file
description: Reads scaled by exon length.

rc_raw

label: Read counts (raw)
type: basic:file
description: Reads uniquely mapped to gene exons.

exp_json

label: Expression RPKUM (polyA) (json)
type: basic:json

exp_type

label: Expression Type (default output)
type: basic:string

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

feature_type

label: Feature type
type: basic:string

Differential Expression (table)¶

data:differentialexpression:uploadupload-diffexp (basic:file src, basic:string gene_id, basic:string logfc, basic:string fdr, basic:string logodds, basic:string fwer, basic:string pvalue, basic:string stat, basic:string source, basic:string species, basic:string build, basic:string feature_type, list:data:expression case, list:data:expression control)[Source: v1.4.1]

Upload Differential Expression table.

src

label: Differential expression file
type: basic:file
description: Differential expression file. Supported file types: *.xls, *.xlsx, *.tab (tab-delimited file), *.diff. DE file must include columns with log2(fold change) and FDR or pval information. DE file must contain header row with column names. Accepts DESeq, DESeq2, edgeR and CuffDiff output files.
validate_regex: \.(xls|xlsx|tab|tab.gz|diff|diff.gz)$

gene_id

label: Gene ID label
type: basic:string

logfc

label: LogFC label
type: basic:string

fdr

label: FDR label
type: basic:string
required: False

logodds

label: LogOdds label
type: basic:string
required: False

fwer

label: FWER label
type: basic:string
required: False

pvalue

label: Pvalue label
type: basic:string
required: False

stat

label: Statistics label
type: basic:string
required: False

source

label

Gene ID database

type

basic:string

choices

AFFY: AFFY
DICTYBASE: DICTYBASE
ENSEMBL: ENSEMBL
NCBI: NCBI
UCSC: UCSC

species

label

Species

type

basic:string

description

Species latin name.

choices

Homo sapiens: Homo sapiens
Mus musculus: Mus musculus
Rattus norvegicus: Rattus norvegicus
Dictyostelium discoideum: Dictyostelium discoideum
Odocoileus virginianus texanus: Odocoileus virginianus texanus
Solanum tuberosum: Solanum tuberosum

build

label: Build
type: basic:string
description: Genome build or annotation version.

feature_type

label

Feature type

type

basic:string

default

gene

choices

gene: gene
transcript: transcript
exon: exon

case

label: Case
type: list:data:expression
description: Case samples (replicates)
required: False

control

label: Control
type: list:data:expression
description: Control samples (replicates)
required: False

raw

label: Differential expression
type: basic:file

de_json

label: Results table (JSON)
type: basic:json

de_file

label: Results table (file)
type: basic:file

source

label: Gene ID database
type: basic:string

species

label: Species
type: basic:string

build

label: Build
type: basic:string

feature_type

label: Feature type
type: basic:string

Differential expression of shRNA¶

data:shrna:differentialexpression:differentialexpression-shrna (data:file parameter_file, list:data:expression:shrna2quant: expression_data)[Source: v1.2.1]

Performing differential expression on a list of objects. Analysis starts by inputting a set of expression files (count matrices) and a parameter file. Parameter file is an xlsx file and consists of tabs: - `sample_key`: Should have column sample with exact sample name as input expression file(s), columns defining treatment and lastly a column which indicates replicate. - `contrasts`: Define groups which will be used to perform differential expression analysis. Model for DE uses these contrasts and replicate number. In R annotation, this would be ` ~ 1