4.5. Variant Calling


Variant calling


fastq file from Illumina Sequencing instrument


vcf and html files

Config file requirements:
  • samples:file1
  • samples:file2
  • project
  • reference:reference.fasta

4.5.1. Details

Snakemake variant calling pipeline based on pipelines of Varun Khanna (https://github.com/khannavarun) and Adrien Villain (https://github.com/avillain). Reads (paired or single) are mapped using bwa mem. Aligned reads are processed with picard tools markduplicates and GATK indel realigner. Freebayes is used to detect SNPs and short INDELs. An annotation file can be set to annotate detected variants. Variants are reported in a HTML report.

The pipeline provides a coverage analysis of the mapping coverage after bam processing. Coverage for each base position is computed with bedtools genomecov. Sequana provides a HTML report with dynamics plots of sequencing coverage and shows interesting regions which have unusual coverage depth.

4.5.2. Usage

sequana --pipeline variant_calling --file1 R1.fastq.gz --file2 R2.fastq.gz --project variant --reference reference.fasta
cd variant
snakemake -s variant_calling.rules -p --stats stats.txt -j 4

4.5.3. Requirements

  • gatk (genomeAnalysisTK)
  • picard (picard-tools)
  • bedtools
  • bwa
  • freebayes
  • snpEff
  • samtools