pileup_reads

Pileup reads from ChIP/ATAC/RNA-seq to make bedgraph/wig/bigwig.

Requirements

• Python >= 3.8, no third-party packages
• samtools on PATH (module load samtools)
• for bigwig, UCSC wigToBigWig / bedGraphToBigWig on PATH; otherwise they are fetched over https and md5-checked into the run's temporary directory. Put them on PATH if compute nodes have no outbound network.

Usage

pileup-reads $INPUT $OUTPUT $OPTIONS...              # installed
python -m pileup_reads $INPUT $OUTPUT $OPTIONS...    # from a checkout

# ChIP
pileup-reads in.bam out.cpm.bigwig -n cpm -p 20
# ATAC
pileup-reads in.bam out.cpm.bigwig -s m36 -t force-extend -e 80 -n cpm -p 20
# RNA-seq
pileup-reads in.bam out.cpm.bigwig -b 1 -t splice -n cpm -p 20
# split a paired-end bam by fragment size
pileup-reads split-by-size in.bam -s 150 -i --plot

pileup-reads --help lists every option. A minus sign may be written m (-s m36 is -36), and counts take a metric prefix (10M).

Notes

• Output does not depend on --chunk-size; tests/ asserts it.
• Unmapped, secondary and supplementary reads are always skipped. Duplicates (0x400) and low MAPQ are only filtered with -f: -f 0 drops duplicates, -f 30 also drops reads below MAPQ 30.
• -t extend builds the interval from the mate, so it also skips reads whose pairing failed (mate unmapped, not properly paired). That is a quality filter: reads the aligner reports as single-end are extended normally.
• .bai and .csi are both found, as file.bam.bai or file.bai.
• Intermediate files go to $SLURM_TMPDIR when set, otherwise $OUTPUT.tmp beside the output, so they land on a filesystem with room. --tmp-dir overrides both.

Tests

module load samtools
python -m unittest discover -s tests -t .   # synthetic fixtures, no data/ needed
tests/smoke.sh --check                      # rebuild data/ tracks and diff
tests/smoke.sh                              # regenerate them in place

data/ holds a real mouse bam and reference bigwigs, kept out of version control for size; tests/ is what guards behaviour. tests/smoke.sh records the settings each reference track was built with.