A Bioinformatics Pipeline for Whole Exome Sequencing: Overview of the Processing and Steps from Raw Data to Downstream Analysis

Abstract Recent advances in next generation sequencing (NGS) technologies have given an impetus to find causality for rare genetic disorders. Since 2005 and aftermath of the human genome project, efforts have been made to understand the rare variants of genetic disorders. Benchmarking the bioinformatics pipeline for whole exome sequencing (WES) has always been a challenge. In this protocol, we discuss detailed steps from quality check to analysis of the variants using a WES pipeline comparing them with reposited public NGS data and survey different techniques, algorithms and software tools used during each step. We observed that variant calling performed on exome and whole genome datasets have different metrics generated when compared to variant callers, GATK and VarScan with different parameters. Furthermore, we found that VarScan with strict parameters could recover 80-85% of high quality GATK SNPs with decreased sensitivity from NGS data. We believe our protocol in the form of pipeline can be used by researchers interested in performing WES analysis for genetic diseases and by large any clinical phenotypes.