Validation of a customized bioinformatics pipeline for a clinical next-generation sequencing test targeting solid tumor-associated variants.

07:00 EST 19th February 2018 | BioPortfolio

Summary of "Validation of a customized bioinformatics pipeline for a clinical next-generation sequencing test targeting solid tumor-associated variants."

Bioinformatic analysis is an integral and critical part of clinical next-generation sequencing. It is especially challenging for some pipelines to consistently identify insertions and deletions. We present the validation of an open source tumor amplicon pipeline (OTA-pipeline) for clinical next-generation sequencing targeting solid tumor-associated variants. Raw data generated from 557 TruSight Tumor 26 samples as well as in silico data were analyzed by the OTA-pipeline and legacy pipeline and compared. Discrepant results were confirmed by orthogonal methods. The OTA-pipeline reported 22 variants which were not detected by the previously validated pipeline, including seven synonymous or intronic single nucleotide variants, five single nucleotide variants at frequency <5%, one insertion, and nine deletions. Variant allele frequencies reported by the two pipelines were highly concordant, although a few significant discrepancies were present. Analysis of in silico FASTQ files demonstrated a higher sensitivity of detecting complex insertions and deletions with the OTA-pipeline. The higher sensitivity came at a cost, as false positive calls were increased in difficult-to-sequence regions. However, these calls were all flagged by our "strand bias" filter, distinguishing them from true variants. Our validation process provides a model for laboratories that wish to establish an in-house bioinformatics pipeline for clinical next-generation sequencing.


Journal Details

This article was published in the following journal.

Name: The Journal of molecular diagnostics : JMD
ISSN: 1943-7811


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