chip-seq-peak-calling-workflow

chip-seq-peak-calling-workflow

Summary

A complete ChIP-Seq peak calling workflow that decomposes MACS3 callpeak into sequential subcommands (filterdup, predictd, pileup, bdgcmp, bdgopt, bdgpeakcall) to progressively transform aligned reads into peak calls with customizable statistical scoring and filtering.

When to use

When you have aligned ChIP-Seq reads (single-end BED or paired-end BEDPE format) and need to identify enriched genomic regions by comparing ChIP signal against control background, with the ability to customize fragment length estimation, local bias calculation, and peak score thresholds rather than using the monolithic callpeak command.

When NOT to use

• Input reads are already in a pre-processed or normalized format (e.g., pre-computed coverage tracks, counts per genomic bin) — the workflow requires raw aligned reads in BED/BEDPE format.
• You need to call broad peaks (e.g., for histone marks covering large domains) — use macs3 bdgbroadcall instead of bdgpeakcall, or apply a different workflow designed for broad mark analysis.
• You lack a suitable control/input sample — the workflow requires both ChIP and control samples to compute local bias; single-sample peak calling requires alternative statistical approaches.

Inputs

• Aligned ChIP-Seq reads in BED format (single-end) or BEDPE format (paired-end)
• Aligned control/input reads in BED format (single-end) or BEDPE format (paired-end)
• Genome size in base pairs (for genome-wide background calculation)
• Sequencing read length (for gap parameter in peak calling)

Outputs

• Filtered ChIP read count (integer)
• Filtered control read count (integer)
• Estimated fragment length d in base pairs (integer)
• ChIP pileup BEDGRAPH file (bedGraph format)
• Local bias BEDGRAPH file (bedGraph format, combined maximum of d/slocal/llocal backgrounds)
• Score BEDGRAPH file (bedGraph format, q-value or p-value scores per base pair)
• narrowPeak file (BED-like format with peak coordinates, summit positions, and score)