Article ID Journal Published Year Pages File Type
3327730 The Journal of Molecular Diagnostics 2015 11 Pages PDF
Abstract

In inherited cardiomyopathies, genetic testing is recognized as an enriching procedure in the diagnostic closure of a cardiac condition. Many genetic mutations have been described as pathogenically related to cardiomyopathies, turning next-generation sequencing into an extremely reliable scenario. Here we describe the validation process of a pipeline constructed with a target panel of 74 cardiomyopathy-related genes sequenced using a next-generation sequencing system. Fifty-two samples from a hypertrophic cardiomyopathy casuistic with previous molecular diagnostics (Sanger-sequenced for MYH7, MYBCP3, and TNNT2; 19 positives and 33 negatives) were processed in parallel with a HapMap reference sample (NA12878) applied for a complete panel assessment. Sequencing coverage values were satisfactory, with a mean of 250× (95% CI, 226.03–273.91) and 95.2% of target bases with a coverage of ≥10×. With a total of 567 variants, variant call sensitivity was tested in five scenarios of coverage and variant allele frequency cutoffs. Maximum achieved sensitivity was 96.7% for single-nucleotide variants and 28.5% for indels, and positive predictive values remained above 0.959 during the whole process. Inter- and intra-assay reproducibility values were 89.5% and 87.3%, respectively. After a careful assessment of analytical performance, we infer that the assay presents potential feasibility for application in diagnostic routines, with minimal time requirements and a simple bioinformatics structure.

Related Topics
Health Sciences Medicine and Dentistry Health Informatics
Authors
, , , , , , ,