atacseq/main
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Tests Complete ATAC-seq analysis pipeline for paired-end reads. Processes raw FASTQ data through adapter and bad quality removal (fastp), alignment (Bowtie2 end-to-end), and filtering (removes MT reads, discordant pairs, low mapping quality below 30, PCR duplicates). Generates 5' cut site pileups (±100bp), performs peak calling, and quantifies reads in 1kb summit-centered regions. Produces two normalized coverage tracks (per million mapped reads and per million reads in peaks) and fragment length distribution plots for quality assessment.
Inputs
| ID | Name | Description | Type |
|---|---|---|---|
| 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 |
|
| PE fastq input | #main/PE fastq input | Should be a paired collection with ATAC-seq fastqs |
|
| Percentage of bad quality bases per read | #main/Percentage of bad quality bases per read | fastp will discard any read which has more than this percentage of bases with a quality below 30, use 100 to not filter reads based on quality. We recommend to use a value that corresponds approximately to 15bp of good quality (for 100bp reads use 85, for 50bp use 70) |
|
| Reference Genome | #main/Reference Genome | Choose a reference genome to map on |
|
Steps
| ID | Name | Description |
|---|---|---|
| 5 | Fastp (remove adapter and bad quality reads) | toolshed.g2.bx.psu.edu/repos/iuc/fastp/fastp/1.3.1+galaxy0 |
| 6 | Bowtie2 map on reference | toolshed.g2.bx.psu.edu/repos/devteam/bowtie2/bowtie2/2.5.5+galaxy0 |
| 7 | filter MAPQ30 concordant pairs and not mitochondrial pairs | toolshed.g2.bx.psu.edu/repos/devteam/bamtools_filter/bamFilter/2.5.3+galaxy0 |
| 8 | Get number of reads per chromosome | toolshed.g2.bx.psu.edu/repos/devteam/samtools_idxstats/samtools_idxstats/2.0.8 |
| 9 | remove PCR duplicates | toolshed.g2.bx.psu.edu/repos/devteam/picard/picard_MarkDuplicates/3.1.1.0 |
| 10 | reads in chrM/MT for multiQC | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/9.5+galaxy3 |
| 11 | convert BAM to BED to improve peak calling | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_bamtobed/2.31.1+galaxy0 |
| 12 | Compute fragment length histogram | toolshed.g2.bx.psu.edu/repos/iuc/pe_histogram/pe_histogram/1.0.2 |
| 13 | number of reads | toolshed.g2.bx.psu.edu/repos/iuc/samtools_view/samtools_view/1.22+galaxy2 |
| 14 | Call Peak with MACS2 | toolshed.g2.bx.psu.edu/repos/iuc/macs2/macs2_callpeak/2.2.9.1+galaxy0 |
| 15 | remove comments lines | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_grep_tool/9.5+galaxy3 |
| 16 | compute 1/million reads | toolshed.g2.bx.psu.edu/repos/devteam/column_maker/Add_a_column1/2.1+galaxy0 |
| 17 | Bigwig from MACS2 (no norm) | wig_to_bigWig |
| 18 | summary of MACS2 | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_grep_tool/9.5+galaxy3 |
| 19 | get summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_slopbed/2.31.1+galaxy0 |
| 20 | Convert 1/million reads to parameter | param_value_from_file |
| 21 | Isolate each bigwig do normalize not average | __APPLY_RULES__ |
| 22 | Merge summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_mergebed/2.31.1+galaxy2 |
| 23 | normalize by million reads | toolshed.g2.bx.psu.edu/repos/bgruening/deeptools_bigwig_average/deeptools_bigwig_average/3.5.4+galaxy0 |
| 24 | Compute coverage on summits +/-500kb | toolshed.g2.bx.psu.edu/repos/iuc/bedtools/bedtools_coveragebed/2.31.1+galaxy0 |
| 25 | number of reads in peaks | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/9.5+galaxy3 |
| 26 | compute 1/million reads in peaks | toolshed.g2.bx.psu.edu/repos/devteam/column_maker/Add_a_column1/2.1+galaxy0 |
| 27 | Combine number of reads in peaks with total number of reads | cat1 |
| 28 | Convert 1/million reads in peaks to parameter | param_value_from_file |
| 29 | reads in peaks multiQC | toolshed.g2.bx.psu.edu/repos/bgruening/text_processing/tp_awk_tool/9.5+galaxy3 |
| 30 | 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/3.5.4+galaxy0 |
| 31 | MultiQC | toolshed.g2.bx.psu.edu/repos/iuc/multiqc/multiqc/1.33+galaxy2 |
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
v0.11071145
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Activity
Views: 21013 Downloads: 46688 Runs: 3
Created: 21st Oct 2022 at 03:01
Last updated: 17th Jan 2023 at 03:01
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