Bibliographic Details
| Title: |
Next-generation sequencing using microfluidic PCR enrichment for molecular autopsy |
| Authors: |
Hariharan Raju, James S. Ware, Jonathan R. Skinner, Paula L. Hedley, Gavin Arno, Donald R. Love, Christian van der Werf, Jacob Tfelt-Hansen, Bo Gregers Winkel, Marta C. Cohen, Xinzhong Li, Shibu John, Sanjay Sharma, Steve Jeffery, Arthur A. M. Wilde, Michael Christiansen, Mary N. Sheppard, Elijah R. Behr |
| Source: |
BMC Cardiovascular Disorders, Vol 19, Iss 1, Pp 1-10 (2019) |
| Publisher Information: |
BMC, 2019. |
| Publication Year: |
2019 |
| Collection: |
LCC:Diseases of the circulatory (Cardiovascular) system |
| Subject Terms: |
Molecular autopsy, Sudden arrhythmic death syndrome, Sudden unexplained death, Inherited cardiac conditions, Next generation sequencing, Diseases of the circulatory (Cardiovascular) system, RC666-701 |
| More Details: |
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. |
| Document Type: |
article |
| File Description: |
electronic resource |
| Language: |
English |
| ISSN: |
1471-2261 |
| Relation: |
http://link.springer.com/article/10.1186/s12872-019-1154-8; https://doaj.org/toc/1471-2261 |
| DOI: |
10.1186/s12872-019-1154-8 |
| Access URL: |
https://doaj.org/article/dcacc9ba179d4d699a6d0ada9f7f32ac |
| Accession Number: |
edsdoj.9ba179d4d699a6d0ada9f7f32ac |
| Database: |
Directory of Open Access Journals |
