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Next-generation sequencing (NGS) provides an unprecedented opportunity to assess genetic variation underlying human disease. Here, we compared two NGS approaches for diagnostic sequencing in inherited arrhythmia syndromes. We compared PCR-based target enrichment and long-read sequencing (PCR-LR) with in-solution hybridization-based enrichment and short-read sequencing (Hyb-SR). The PCR-LR assay comprehensively assessed five long-QT genes routinely sequenced in diagnostic laboratories and "hot spots" in RYR2. The Hyb-SR assay targeted 49 genes, including those in the PCR-LR assay. The sensitivity for detection of control variants did not differ between approaches. In both assays, the major limitation was upstream target capture, particular in regions of extreme GC content. These initial experiences with NGS cardiovascular diagnostics achieved up to 89 % sensitivity at a fraction of current costs. In the next iteration of these assays we anticipate sensitivity above 97 % for all LQT genes. NGS assays will soon replace conventional sequencing for LQT diagnostics and molecular pathology.

Original publication

DOI

10.1007/s12265-012-9401-8

Type

Journal article

Journal

Journal of cardiovascular translational research

Publication Date

02/2013

Volume

6

Pages

94 - 103

Addresses

MRC Clinical Sciences Centre, Imperial College London, London, UK. j.ware@imperial.ac.uk

Keywords

Humans, Genetic Predisposition to Disease, Ion Channels, Ryanodine Receptor Calcium Release Channel, Potassium Channels, Voltage-Gated, Genetic Markers, Sensitivity and Specificity, Predictive Value of Tests, Polymerase Chain Reaction, DNA Mutational Analysis, Heredity, Mutation, Ether-A-Go-Go Potassium Channels, KCNQ1 Potassium Channel, Arrhythmias, Cardiac, Genetic Testing, High-Throughput Nucleotide Sequencing, NAV1.5 Voltage-Gated Sodium Channel, ERG1 Potassium Channel