Version Compatibility

Reference examples tested with: Bowtie2 2.5.3+, MACS3 3.0+, samtools 1.19+

Before using code patterns, verify installed versions match. If versions differ:

• CLI: <tool> --version then <tool> --help to confirm flags

If code throws ImportError, AttributeError, or TypeError, introspect the installed package and adapt the example to match the actual API rather than retrying.

ATAC-seq Peak Calling

"Call peaks from my ATAC-seq data" → Identify open chromatin regions using ATAC-specific parameters (no input control, shifted Tn5 cut sites, paired-end mode).

• CLI: macs3 callpeak -t atac.bam -f BAMPE -g hs --nomodel --shift -75 --extsize 150

Basic MACS3 for ATAC-seq

Goal: Identify open chromatin regions from ATAC-seq data using ATAC-specific peak calling parameters.

Approach: Run MACS3 in paired-end mode with Tn5 shift correction, no model building, and duplicate retention since ATAC-seq generates natural duplicates at accessible sites.

# Standard ATAC-seq peak calling
macs3 callpeak \
    -t sample.bam \
    -f BAMPE \
    -g hs \
    -n sample \
    --outdir peaks/ \
    -q 0.05 \
    --nomodel \
    --shift -75 \
    --extsize 150 \
    --keep-dup all \
    -B

Key ATAC-seq Parameters

# Explained parameters
macs3 callpeak \
    -t sample.bam \        # Treatment BAM
    -f BAMPE \             # Paired-end BAM (uses fragment size)
    -g hs \                # Genome size: hs (human), mm (mouse)
    -n sample \            # Output name prefix
    --nomodel \            # Don't build shifting model
    --shift -75 \          # Shift reads to center on Tn5 cut site
    --extsize 150 \        # Extend reads to this size
    --keep-dup all \       # Keep duplicates (ATAC has natural duplicates)
    -B \                   # Generate bedGraph for visualization
    --call-summits         # Call peak summits

Why These Parameters?