RNAseq-variant-calling-nf

RNAseq-variant_calling-nf is a A Nextflow workflow to call variants with GATK from bulk RNAseq data

Usage

nextflow run RNAseq-variant-calling-nf --fastq [path to fastqs] --project [project ID]

Required arguments:

• --fastq
  • Path to fastqs, default is ./Fastq
• --project
  • Project name. E.g., '19482R'. Will be given to final vcf file

Outline

This workflow will:

• Make channel from paired end reads
• Remove optical duplicates with Clumpify
• Trim adapters with Cutadapt (hardcoded adapter sequence)
• Map with STAR. BAMs published to outdir/bams
• Index BAMS and calculate idxstats with Samtools. Also published to oudir/bams
• featureCounts. Published to outdir/feature_counts
• RSEM gene and isoform counts are published to outdir/rsem
• RNAseq metrics with Picard. Published to outdir/rnaseq_metrics
• Fliter alignments for unmapped, secondary, qcfail, and supplementary
• Mark duplicates with Picard and reindex.
• Generate read coverage with USeq and convert to BigWig. Published to outdir/coverage
• Split and trim intron junctions
• Indel realigner
• Base recalibration
• Pad BED file
• Haplotype variant calls. Published to outdir/haplotype_vcf
• Variant filtering
• Merge raw variants
• Generate genomic VCF for joint genotyping
• Joint genotyping. Published to outdir/final

Results

Workflow results are published to --outdir, whose default is ./results
These include:

• BAMs from STAR to ./results/bams
• Samtools idxstats to ./results/bams
• featureCounts to ./results/feature_counts
• RSEM to ./results/rsem
• RNAseq metrics to ./results/rnaseq_metrics
• BigWigs to ./results/coverage
• Per sample GATK haplotype calls to ./results/haplotype_vcf
• Joint genotyping with GATK to ./results/final

Defaults

Default values for paths to applications and reference files are in nextflow.config.
Default organism is human with Ensembl v96 annotation