/pipeline-chipseq

name: pipeline-chipseq
description: "Execute ENCODE ChIP-seq processing pipeline from FASTQ to peaks and signal tracks. Child of pipeline-guide. Provides stage-by-stage Nextflow execution with Docker containers and cloud deployment. Use when users need to process ChIP-seq data following ENCODE standards, run peak calling with MACS2, perform IDR analysis, or generate signal tracks. Trigger on: ChIP-seq pipeline, run ChIP-seq, process ChIP-seq, MACS2 peak calling, IDR analysis, ChIP-seq FASTQ processing."

ENCODE ChIP-seq Pipeline

When to Use

• User wants to run a ChIP-seq processing pipeline from FASTQ to peaks and signal tracks
• User asks about "ChIP-seq pipeline", "MACS2", "peak calling", "BWA alignment for ChIP", or "IDR"
• User needs to process histone or TF ChIP-seq data following ENCODE standards
• Example queries: "process my ChIP-seq FASTQs", "run the ENCODE ChIP-seq pipeline", "call peaks from ChIP-seq with MACS2 and IDR"

Execute the ENCODE ChIP-seq processing pipeline from raw FASTQ files through peak calling, IDR analysis, and signal track generation. This skill provides a Nextflow DSL2 implementation following ENCODE uniform analysis standards.

Overview

The pipeline processes chromatin immunoprecipitation sequencing data through quality control, adapter trimming, alignment to a reference genome, filtering and duplicate removal, blacklist filtering, peak calling with MACS2, an IDR comparison for every pair of replicates, FRiP calculation, and signal track generation.

The same workflow handles transcription factor (TF) ChIP-seq and histone modification ChIP-seq. The peak mode is chosen once per run with `--peak_type narrow|broad`; it applies to every sample in that run. To process narrow and broad targets together, run the workflow twice with different `--peak_type` and `--outdir` values.

Key Literature

| Reference | Journal | Year | DOI | Relevance | |-----------|---------|------|-----|-----------| | Landt et al. "ChIP-seq guidelines and practices" | Genome Research | 2012 | 10.1101/gr.136184.111 | ENCODE ChIP-seq standards (~4,000 citations) | | ENCODE Project Consortium "Expanded encyclopaedias" | Nature | 2020 | 10.1038/s41586-020-2493-4 | ENCODE Phase 3 standards | | Zhang et al. "Model-based Analysis of ChIP-Seq (MACS)" | Genome Biology | 2008 | 10.1186/gb-2008-9-9-r137 | Peak caller (~7,000 citations) | | Li et al. "Measuring reproducibility (IDR)" | Annals of Applied Statistics | 2011 | 10.1214/11-AOAS466 | Replicate consistency (~1,500 citations) | | Amemiya et al. "ENCODE Blacklist" | Scientific Reports | 2019 | 10.1038/s41598-019-45839-z | Artifact regions (~1,372 citations) | | Ramachandran et al. "phantompeakqualtools" | — | 2013 | — | NSC/RSC strand correlation metrics (manual step, see below) |

Pipeline Stages

FASTQ ──> FastQC / Trim Galore ──> BWA-MEM ──> Samtools Filter ──> Picard MarkDuplicates
  │                                            (-F 1804/-F 1028, -q 30)   (duplicates REMOVED)
  │                                                                       │
  │           ┌───────────────────────────────────────────────────────────┘
  │           v
  │     Blacklist Filter ──> MACS2 Peak Calling ──> IDR (narrow runs only, every pair)
  │       (applied to BAM)          │
  │                                 ├──> Signal Tracks (bdgcmp + bedGraphToBigWig)
  │                                 └──> FRiP (bedtools intersect + samtools view -c)
  v
 QC reports ──────────────────────────────────> MultiQC

Control libraries, when `--control` is given, travel through stages 1-3 alongside the ChIP samples with a `CONTROL_` prefix and are split off before peak calling; MACS2 pools them with `-c`.

Stage Summary

| Stage | Tool | Input | Output | Reference | |-------|------|-------|--------|-----------| | 1. QC & Trimming | FastQC, Trim Galore | Raw FASTQ | Trimmed FASTQ, FastQC reports | references/01-qc-trimming.md | | 2. Alignment | BWA-MEM, samtools | Trimmed FASTQ | Sorted BAM, flagstat | references/02-alignment.md | | 3. Filtering | samtools, Picard, bedtools | Sorted BAM | Deduplicated, blacklist-filtered BAM | references/03-filtering.md | | 4. Peak Calling & IDR | MACS2, IDR | Filtered BAM | narrowPeak/broadPeak, one `<sampleA>_vs_<sampleB>.idr_peaks.txt` per replicate pair | references/04-analysis.md | | 5. Signal, FRiP & QC report | MACS2 bdgcmp, bedGraphToBigWig, bedtools, samtools, MultiQC | MACS2 bedGraphs, final BAM, peaks, QC logs | bigWig, `<sample>.frip_mqc.tsv`, multiqc_report.html | references/05-qc-metrics.md |

Input Requirements

Required

• **Treatment FASTQ** (`--reads`): ChIP sample reads, gzipped. A Nextflow file-pair glob,

e.g. `'fastq/chip_*_R{1,2}.fq.gz'`. Paired-end by default; add `--single_end` for SE data.

• **Chromosome sizes** (`--chrom_sizes`): two-column `<chrom>\t<size>` file used by

`bedGraphToBigWig`. The workflow stops immediately if it is missing. Build it with `samtools faidx GRCh38.fa && cut -f1,2 GRCh38.fa.fai > GRCh38.chrom.sizes`.

• **BWA index directory** (`--bwa_index`): a directory holding `<genome>.fa` plus its BWA

index files. BWA is invoked as `bwa mem ... <dir>/GRCh38.fa`, so the directory must contain `GRCh38.fa`, `GRCh38.fa.amb`, `GRCh38.fa.ann`, `GRCh38.fa.bwt`, `GRCh38.fa.pac` and `GRCh38.fa.sa`. Build it once with `bwa index GRCh38_index/GRCh38.fa`. If the flag is omitted, the workflow looks for `./<genome>_index` in the launch directory. The workflow does not build or download the index.

Optional

• **Control FASTQ** (`--control`): input/IgG reads, same glob form. Strongly recommended

for meaningful enrichment, but the workflow runs without one; MACS2 then uses its local lambda background model.

• **Blacklist** (`--blacklist`): defaults to the ENCODE Blacklist v2 URL for `--genome`.

There is no sample sheet. Inputs are globs, and every sample in a run shares one `--peak_type` and one `--genome`.

**The `--reads` and `--control` globs mus