Abstract
BackgroundComplex insertions and deletions (indels) from next-generation sequencing (NGS) data were prone to escape detection by currently available variant callers as shown by large-scale human genomics studies. Somatic and germline complex indels in key disease driver genes could be missed in NGS-based genomics studies.ResultsINDELseek is an open-source complex indel caller designed for NGS data of random fragments and PCR amplicons. The key differentiating factor of INDELseek is that each NGS read alignment was examined as a whole instead of “pileup” of each reference position across multiple alignments. In benchmarking against the reference material NA12878 genome (n = 160 derived from high-confidence variant calls), GATK, SAMtools and INDELseek showed complex indel detection sensitivities of 0%, 0% and 100%, respectively. INDELseek also detected all known germline (BRCA1 and BRCA2) and somatic (CALR and JAK2) complex indels in human clinical samples (n = 8). Further experiments validated all 10 detected KIT complex indels in a discovery cohort of clinical samples. In silico semi-simulation showed sensitivities of 93.7–96.2% based on 8671 unique complex indels in >5000 genes from dbSNP and COSMIC. We also demonstrated the importance of complex indel detection in accurately annotating BRCA1, BRCA2 and TP53 mutations with gained or rescued protein-truncating effects.ConclusionsINDELseek is an accurate and versatile tool for complex indel detection in NGS data. It complements other variant callers in NGS-based genomics studies targeting a wide spectrum of genetic variations.
Highlights
Complex insertions and deletions from next-generation sequencing (NGS) data were prone to escape detection by currently available variant callers as shown by large-scale human genomics studies
We observed closely spaced single nucleotide variant (SNV) that appeared in trans in the alignments and 26 such loci were manually curated as negative controls for complex indel detection (Additional file 1: Table S2)
To demonstrate the importance of accurate complex indel detection in clinical settings, we focused on 127 multiple-nucleotide variants (MNV) in hereditary breast and/or ovarian cancer (HBOC) genes and compared their variant annotation results (Variant Effect Predictor) in two scenarios: (1) original MNV and (2) decomposing MNV into individual single-nucleotide variant for separate annotation, as if the MNV could not be called as a haplotype
Summary
Complex insertions and deletions (indels) from next-generation sequencing (NGS) data were prone to escape detection by currently available variant callers as shown by large-scale human genomics studies. Somatic and germline complex indels in key disease driver genes could be missed in NGS-based genomics studies. Complex insertions and deletions (indels) are a known class of genetic variation [1] associated with human diseases [2]. Simultaneous deletion and insertion of DNA fragments of different sizes lead to net change in length. No net change in length is possible in case of contiguous or non-contiguous multiple-nucleotide variants (MNV). Recent studies revealed the shortcomings of state-of-the-art variant callers that might fail to detect somatic and germline complex indels [3, 4]. Important mutations in key disease driver genes
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