INDELseek: detection of complex insertions and deletions from next-generation sequencing data

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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3449-9) contains supplementary material, which is available to authorized users.

Next-generation sequencing and molecular cytogenetic characterization of ETV6-LYN fusion due to chromosomes 1, 8 and 12 rearrangement in acute myeloid leukemia

“…The RUNX1T1 signal was detected on the short arm of der(1), while the ETV6 signal was split between der(12) and the tip of the short arm of der(1). The 3′ MYC signal remained on der(8) but 5′ MYC signal was not detected on der(1) or der (8). While the split MYC signal further confirmed the breakpoint on chromosome 8, a bona fide MYC rearrangement in this patient could not be excluded.…”

“…Briefly, 50 ng extracted DNA was subject to PCR amplification using Illumina TruSight Myeloid panel (568 amplicons for 54 genes) and followed by paired-end MiSeq sequencing (2 × 275bp). Called single-nucleotide variants, small insertions/deletions (indels), complex indels (8), and FLT3 internal tandem duplication variants (minimum position sequencing depth 500-fold and minimum variant allele fraction 0.05) were manually examined using Integrative Genomics Viewer (IGV) version 2.1.30. Any variants reported by 1000 Genomes Project Phase 3 were excluded.…”

“…More recently, ETV6-LYN gene fusion was detected in a patient who suffered from MPN with eosinophilia and T-lymphoblastic lymphoma, fitting within the category of myeloid/lymphoid neoplasms associated with eosinophilia (3). The gene fusion occurred in the form of der (8)inv(q12.1q21.1)t(8;12)(q12.1;p13) postulated to involve at least three breaks. The simplest form involved an inversion of chromosome 8q and fusion with chromosome 12p13, which brought ETV6 and LYN into juxtaposition in the correct 5′ to 3′ orientation to create the in-frame chimeric fusion gene on the derivative chromosome 8 that was necessary owing to the opposite orientation of the genes on the respective chromosomes (3).…”

Analysis and Annotation of Whole‐Genome or Whole‐Exome Sequencing Derived Variants for Clinical Diagnosis

“…This is particularly true for identification of complex indels that may have arisen through multiple insertion, deletion, and substitution events (Au, Leung, Kwong, Chan, & Ma, ; Fang et al., ; Ye et al., ). As might therefore be expected, estimates of concordance among commonly used IDI callers are lower than for SNVs, though more recent reports depict increased sensitivity and specificity when attempting to re‐identify known variants from control datasets (Au et al., ; Hofmann et al., ). This remains an area of active methods development (e.g., Au et al., ; Fang et al., ; Lek et al., ; Ye et al., ).…”

“…There is ongoing debate as to the comparative sensitivity and selectivity of the various IDI callers; it is acknowledged that none of the existing tools is unequivocally best for all cases (e.g., Au et al., ; Fang et al., ; Lek et al., ; O'Rawe et al., ; Ye et al., ). Of the many IDI calling tools available, the GATK package from the Broad Institute remains the selection at many institutions, likely in part due to the fact that it can be used for both IDI and SNV calling (McKenna et al., ).…”