We have developed and validated a clinical whole-genome and transcriptome sequencing (WGTS) assay which provides a comprehensive genomic profile of a patient’s tumor. The ability to fully capture the mappable genome with sufficient sequencing coverage to precisely call DNA somatic single nucleotide variants, Indels, copy number variants, structural variants, and RNA gene fusions, was analyzed. New York State’s Department of Health NGS guidelines were expanded on for establishing performance validation applicable to whole-genome sequencing. Whole-genome sequencing laboratory protocols were validated for the Illumina HiSeq X Ten platform and RNA sequencing for Illumina HiSeq2500 platform for fresh frozen and formalin-fixed, paraffin-embedded tumor samples. Various bioinformatics tools were also tested, and confidence intervals for sensitivity and specificity thresholds in calling clinically significant somatic aberrations were determined. The validation was performed on a set of 125 tumor normal pairs. RNA sequencing is performed to call fusion and to confirm the DNA variants or exonic alterations. Here, we present our results and WGTS standards for variant allele frequency, reproducibility, analytical sensitivity, and present limit of detection analysis for single nucleotide variant calling, copy number identification, and structural variants. We show that The New York Genome Center WGTS clinical assay can provide a comprehensive patient variant discovery approach suitable for directed oncological therapeutic applications.

PMID: 30138725 DOI: 10.1016/j.jmoldx.2018.06.007

J Mol Diagn. 2018 Aug 20. pii: S1525-1578(17)30621-9. doi: 10.1016/j.jmoldx.2018.06.007.