Next-generation sequencing using microfluidic PCR enrichment for molecular autopsy.

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Show simple item record Raju, Hariharan en Ware, James S en Skinner, Jonathan en Hedley, Paula L en Arno, Gavin en Love, Donald en van der Werf, Christian en Tfelt-Hansen, Jacob en Winkel, Bo Gregers en Cohen, Marta C en Li, Xinzhong en John, Shibu en Sharma, Sanjay en Jeffery, Steve en Wilde, Arthur AM en Christiansen, Michael en Sheppard, Mary N en Behr, Elijah R en 2020-02-07T00:55:38Z en 2019-07-23 en
dc.identifier.citation BMC cardiovascular disorders 19(1):174 23 Jul 2019 en
dc.identifier.issn 1471-2261 en
dc.identifier.uri en
dc.description.abstract BACKGROUND:We aimed to determine the mutation yield and clinical applicability of "molecular autopsy" following sudden arrhythmic death syndrome (SADS) by validating and utilizing low-cost high-throughput technologies: Fluidigm Access Array PCR-enrichment with Illumina HiSeq 2000 next generation sequencing (NGS). METHODS:We validated and optimized the NGS platform with a subset of 46 patients by comparison with Sanger sequencing of coding exons of major arrhythmia risk-genes (KCNQ1, KCNH2, SCN5A, KCNE1, KCNE2, RYR2). A combined large multi-ethnic international SADS cohort was sequenced utilizing the NGS platform to determine overall molecular yield; rare variants identified by NGS were subsequently reconfirmed by Sanger sequencing. RESULTS:The NGS platform demonstrated 100% sensitivity for pathogenic variants as well as 87.20% sensitivity and 99.99% specificity for all substitutions (optimization subset, n = 46). The positive predictive value (PPV) for NGS for rare substitutions was 16.0% (27 confirmed rare variants of 169 positive NGS calls in 151 additional cases). The overall molecular yield in 197 multi-ethnic SADS cases (mean age 22.6 ± 14.4 years, 68% male) was 5.1% (95% confidence interval 2.0-8.1%), representing 10 cases carrying pathogenic or likely pathogenic risk-mutations. CONCLUSIONS:Molecular autopsy with Fluidigm Access Array and Illumina HiSeq NGS utilizing a selected panel of LQTS/BrS and CPVT risk-genes offers moderate diagnostic yield, albeit requiring confirmatory Sanger-sequencing of mutational variants. en
dc.format.medium Electronic en
dc.language eng en
dc.relation.ispartofseries BMC cardiovascular disorders en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. en
dc.rights.uri en
dc.rights.uri en
dc.subject Humans en
dc.subject Death, Sudden, Cardiac en
dc.subject Genetic Predisposition to Disease en
dc.subject Autopsy en
dc.subject Microfluidic Analytical Techniques en
dc.subject Cause of Death en
dc.subject Risk Factors en
dc.subject Reproducibility of Results en
dc.subject Predictive Value of Tests en
dc.subject Polymerase Chain Reaction en
dc.subject Pedigree en
dc.subject DNA Mutational Analysis en
dc.subject Heredity en
dc.subject Mutation en
dc.subject Adolescent en
dc.subject Adult en
dc.subject Child en
dc.subject Child, Preschool en
dc.subject Infant en
dc.subject Australia en
dc.subject Europe en
dc.subject New Zealand en
dc.subject Female en
dc.subject Male en
dc.subject Arrhythmias, Cardiac en
dc.subject Young Adult en
dc.subject Pathology, Molecular en
dc.subject High-Throughput Nucleotide Sequencing en
dc.title Next-generation sequencing using microfluidic PCR enrichment for molecular autopsy. en
dc.type Journal Article en
dc.identifier.doi 10.1186/s12872-019-1154-8 en
pubs.issue 1 en
pubs.begin-page 174 en
pubs.volume 19 en
dc.rights.holder Copyright: The authors en
pubs.publication-status Published en
dc.rights.accessrights en
pubs.subtype Research Support, Non-U.S. Gov't en
pubs.subtype research-article en
pubs.subtype Validation Study en
pubs.subtype Multicenter Study en
pubs.subtype Journal Article en
pubs.elements-id 779007 en
dc.identifier.eissn 1471-2261 en
pubs.record-created-at-source-date 2019-07-25 en
pubs.dimensions-id 31337358 en

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