🧬 chipcallvar

chipcallvar is a reproducible Nextflow workflow for calling somatic variants (SNVs and INDELs) from ChIP-seq data. It supports multiple variant calling tools and is designed for parallel execution, high customizability, and scalability on HPC or cloud environments.

The pipeline integrates the following tools:

1. MACS3 callvar: peak-aware variant caller optimized for ChIP-seq data
2. GATK Mutect2: industry-standard somatic SNV/INDEL caller
3. FreeBayes: haplotype-based variant detection

Workflow Overview

• Read alignment – bwa-mem2, SamTools
• Merging technical replicates – SamTools merge
• Marking duplicate reads - GATK MarkDuplicates
• Base Quality Scores Recalibration - GATK BaseRecalibrator, GATK ApplyBQSR
• Interval creation – BedTools
• Peak calling – MAC3 CallPeak
• Variant calling – MAC3 CallVar, GATK Mutect2, FreeBayes
• Variant annotation – Ensembl VEP
• Filtering and reheadering – BcfTools
• Optional MAF conversion – vcf2maf, maftools
• Quality control and reporting – FastQC, SamTools, Mosdepth, BcfTools, Ensembl VEP, summarized with MultiQC

To speed up variant calling, the reference genome is split into smaller genomic intervals using bedtools. Each interval is processed in parallel, and the resulting VCF files are merged. This significantly reduces the total wall-clock time and optimizes the use of compute resources.

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📥 Input

Sample Sheet

Each row describes one replicate of a ChIP-seq experiment. The input format is:

id,sample,replicate,fastq_1,fastq_2,control,control_replicate
OCI-AML3,OCI-AML3_input,1,test/hs_ChIP_OCI-AML3_rep1_Input_R1_001.fastq.gz, test/hs_ChIP_OCI-AML3_rep1_Input_R2_001.fastq.gz,,
OCI-AML3,OCI-AML3_H3K27ac,1,test/hs_ChIP_OCI-AML3_rep1_H3K27ac_R1_001.fastq.gz,test/hs_ChIP_OCI-AML3_rep1_H3K27ac_R2_001.fastq.gz,OCI-AML3_input,1
OCI-AML3,OCI-AML3_H3K27ac,2,test/hs_ChIP_OCI-AML3_rep2_H3K27ac_R1_001.fastq.gz,test/hs_ChIP_OCI-AML3_rep2_H3K27ac_R2_001.fastq.gz,OCI-AML3_input,1

id,sample,vcf,caller
24_F,SA10,SA10_peaks.vcf,macs3
85_M,SA11,SA11_peaks.vcf,macs3

⚙️ Parameters File

step can be 'mapping', 'variant_calling', 'annotate'

outdir: './results'
samplesheet: './samplesheet_example.csv'
# reference genome
fasta: './resources-broad-hg38-v0-Homo_sapiens_assembly38.fasta'
fasta_fai: './resources-broad-hg38-v0-Homo_sapiens_assembly38.fasta.fai'
assembly: 'GRCh38'
step: 'mapping'
tools: 'macs3,mutect2,freebayes'
skip_tools: 'dge,markduplicates,bqsr'
no_intervals: TRUE
merge_vcfs: TRUE
# variant filtering
depth: 10
vaf: 0.1
af1: 0.0001
af2: 0.0001
# for DGE analysis
tpm: './nf-rnaseq/star_rsem/rsem.merged.gene_tpm.tsv'
email: 'example@gmail.com'

🚀 Running the Pipeline

nextflow run main.nf -params-file params.yaml 

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👩‍💻 Author

Ann Mariya GitHub Email