bio-workflows-chipseq-pipeline

name	bio-workflows-chipseq-pipeline
description	End-to-end ChIP-seq workflow from FASTQ files to annotated peaks. Covers QC, alignment, peak calling with MACS3 (or HOMER), and peak annotation with ChIPseeker. Use when processing ChIP-seq data from alignment through peak annotation.
tool_type	mixed
primary_tool	MACS3
workflow	true
depends_on	["read-qc/fastp-workflow","read-alignment/bowtie2-alignment","alignment-files/duplicate-handling","chip-seq/chipseq-qc","chip-seq/peak-calling","chip-seq/peak-annotation","chip-seq/differential-binding","chip-seq/chipseq-visualization","chip-seq/motif-analysis"]
qc_checkpoints	[{"after_qc":"Q30 >85%, adapter content <5%"},{"after_alignment":"Mapping rate >80%, unique mapping >70%"},{"after_dedup":"NRF >0.8, PBC1 >0.8 (compute pre-dedup)"},{"after_peaks":"FRiP >1% (TF) or >5% (histone); NSC >1.05; RSC >0.8"},{"after_idr":"Nself and Nt ratios both <=2 (ENCODE consistency rule)"}]

Version Compatibility

Reference examples tested with: Bowtie2 2.5.3+, MACS3 3.0+, HOMER 4.11+, bedtools 2.31+, fastp 0.23+, samtools 1.19+

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

• R: packageVersion('<pkg>') then ?function_name to verify parameters
• 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.

ChIP-seq Pipeline

"Process my ChIP-seq data from FASTQ to annotated peaks" → Orchestrate QC, Bowtie2 alignment, duplicate removal, MACS3 peak calling, ChIPseeker annotation, and QC metrics (FRiP, strand cross-correlation).

Complete workflow from raw ChIP-seq FASTQ files to annotated peaks.

Workflow Overview

FASTQ files (IP + Input)
    |
    v
[1. QC & Trimming] -----> fastp
    |
    v
[2. Alignment] ---------> Bowtie2
    |
    v
[3. BAM Processing] ----> sort, markdup, filter
    |
    v
[4. Peak Calling] ------> MACS3
    |
    v
[5. QC] ----------------> FRiP, fingerprint plots
    |
    v
[6. Annotation] --------> ChIPseeker
    |
    v
Annotated peaks + QC report

Primary Path: Bowtie2 + MACS3 + ChIPseeker

Step 1: Quality Control with fastp

# Process both IP and Input samples
for sample in IP_rep1 IP_rep2 Input_rep1 Input_rep2; do
    fastp -i ${sample}_R1.fastq.gz -I ${sample}_R2.fastq.gz \
        -o trimmed/${sample}_R1.fq.gz -O trimmed/${sample}_R2.fq.gz \
        --detect_adapter_for_pe \
        --qualified_quality_phred 20 \
        --length_required 25 \
        --html qc/${sample}_fastp.html
done

Step 2: Alignment with Bowtie2

# Build index (once)
bowtie2-build genome.fa bt2_index/genome

# Align
for sample in IP_rep1 IP_rep2 Input_rep1 Input_rep2; do
    bowtie2 -p 8 -x bt2_index/genome \
        -1 trimmed/${sample}_R1.fq.gz \
        -2 trimmed/_R2.fq.gz \
        --no-mixed --no-discordant \
        --maxins 1000 \
        2> aligned/. | \
    samtools view -@ 4 -bS -q 30 - | \
    samtools  -@ 4 -o aligned/.bam