This workflow takes as input a collection of paired fastq. It will remove bad quality and adapters with cutadapt. Map with Bowtie2 end-to-end. Will remove reads on MT and unconcordant pairs and pairs with mapping quality below 30 and PCR duplicates. Will compute the pile-up on 5' +- 100bp. Will call peaks and count the number of reads falling in the 1kb region centered on the summit. Will compute 2 normalization for coverage: normalized by million reads and normalized by million reads in peaks. Will plot the number of reads for each fragment length.
Inputs
ID | Name | Description | Type |
---|---|---|---|
PE fastq input | #main/PE fastq input | Should be a paired collection with ATAC-seq fastqs |
|
bin_size | #main/bin_size | Bin size for normalized bigwig (usually 50bp is sufficient) |
|
effective_genome_size | #main/effective_genome_size | Used by macs2:\nH. sapiens: 2700000000, M. musculus: 1870000000, D. melanogaster: 120000000, C. elegans: 90000000 |
|
reference_genome | #main/reference_genome | reference_genome |
|
Steps
ID | Name | Description |
---|---|---|
4 | Cutadapt (remove adapter + bad quality bases) | toolshed.g2.bx.psu.edu/repos/lparsons/cutadapt/cutadapt/4.4+galaxy0 |
5 | Bowtie2 map on reference | toolshed.g2.bx.psu.edu/repos/devteam/bowtie2/bowtie2/2.5.0+galaxy0 |
6 | filter MAPQ30 concordant pairs and not mitochondrial pairs | toolshed.g2.bx.psu.edu/repos/devteam/bamtools_filter/bamFilter/2.5.2+galaxy1 |
7 | Get number of reads per chromosome | toolshed.g2.bx.psu.edu/repos/devteam/samtools_idxstats/samtools_idxstats/2.0.4 |
8 | remove PCR duplicates | toolshed.g2.bx.psu.edu/repos/devteam/picard/picard_MarkDuplicates/2.18.2.4 |
9 | reads in chrM/MT for multiQC | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/1.1.2 |
10 | convert BAM to BED to improve peak calling | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_bamtobed/2.30.0+galaxy2 |
11 | Compute fragment length histogram | toolshed.g2.bx.psu.edu/repos/iuc/pe_histogram/pe_histogram/1.0.1 |
12 | number of reads | toolshed.g2.bx.psu.edu/repos/iuc/samtools_view/samtools_view/1.15.1+galaxy0 |
13 | Call Peak with MACS2 | toolshed.g2.bx.psu.edu/repos/iuc/macs2/macs2_callpeak/2.2.7.1+galaxy0 |
14 | compute 1/million reads | toolshed.g2.bx.psu.edu/repos/devteam/column_maker/Add_a_column1/2.0 |
15 | Bigwig from MACS2 (no norm) | wig_to_bigWig |
16 | get summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_slopbed/2.30.0+galaxy1 |
17 | summary of MACS2 | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_grep_tool/1.1.1 |
18 | Convert 1/million reads to parameter | param_value_from_file |
19 | Isolate each bigwig do normalize not average | __APPLY_RULES__ |
20 | Merge summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_mergebed/2.30.0 |
21 | normalize by million reads | toolshed.g2.bx.psu.edu/repos/bgruening/deeptools_bigwig_average/deeptools_bigwig_average/3.5.2+galaxy0 |
22 | Compute coverage on summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_coveragebed/2.30.0+galaxy1 |
23 | number of reads in peaks | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/1.1.2 |
24 | compute 1/million reads in peaks | toolshed.g2.bx.psu.edu/repos/devteam/column_maker/Add_a_column1/2.0 |
25 | Combine number of reads in peaks with total number of reads | cat1 |
26 | Convert 1/million reads in peaks to parameter | param_value_from_file |
27 | reads in peaks multiQC | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/1.1.2 |
28 | normalize by million reads in peaks | Isolate each bigwig do normalize not average toolshed.g2.bx.psu.edu/repos/bgruening/deeptools_bigwig_average/deeptools_bigwig_average/3.5.2+galaxy0 |
29 | MultiQC | toolshed.g2.bx.psu.edu/repos/iuc/multiqc/multiqc/1.11+galaxy1 |
Outputs
ID | Name | Description | Type |
---|---|---|---|
1kb around summits | #main/1kb around summits | n/a |
|
BAM filtered rmDup | #main/BAM filtered rmDup | n/a |
|
Coverage from MACS2 (bigwig) | #main/Coverage from MACS2 (bigwig) | n/a |
|
MACS2 narrowPeak | #main/MACS2 narrowPeak | n/a |
|
MACS2 report | #main/MACS2 report | n/a |
|
MarkDuplicates metrics | #main/MarkDuplicates metrics | n/a |
|
MultiQC on input dataset(s): Stats | #main/MultiQC on input dataset(s): Stats | n/a |
|
MultiQC webpage | #main/MultiQC webpage | n/a |
|
Nb of reads in summits +-500bp | #main/Nb of reads in summits +-500bp | n/a |
|
bigwig_norm | #main/bigwig_norm | n/a |
|
bigwig_norm2 | #main/bigwig_norm2 | n/a |
|
histogram of fragment length | #main/histogram of fragment length | n/a |
|
mapping stats | #main/mapping stats | n/a |
|
Version History
v0.1 (earliest) Created 21st Oct 2022 at 03:01 by WorkflowHub Bot
Updated to v0.1
Frozen
v0.1
1071145
Creator
Additional credit
Lucille Delisle
Submitter
Views: 7815 Downloads: 2188 Runs: 0
Created: 21st Oct 2022 at 03:01
Last updated: 17th Jan 2023 at 03:01
None