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Validation and Optimization of the Ion Torrent S5 XL Sequencer and Oncomine Workflow for BRCA1 and BRCA2 Genetic Testing

 Saeam Shin  ;  Yoonjung Kim  ;  Seoung Chul Oh  ;  Nae Yu  ;  Seung-Tae Lee  ;  Jong Rak Choi  ;  Kyung-A Lee 
 ONCOTARGET , Vol.8(21) : 34858-34866, 2017 
Journal Title
Issue Date
BRCA1 Protein/genetics* ; BRCA2 Protein/genetics* ; DNA Copy Number Variations ; Female ; Genetic Testing/instrumentation* ; Hereditary Breast and Ovarian Cancer Syndrome/genetics* ; High-Throughput Nucleotide Sequencing/instrumentation ; Humans ; INDEL Mutation ; Polymorphism, Single Nucleotide ; Sequence Analysis, DNA/instrumentation
BRCA ; Ion Torrent ; Oncomine ; S5 XL ; next-generation sequencing
In this study, we validated the analytical performance of BRCA1/2 sequencing using Ion Torrent's new bench-top sequencer with amplicon panel with optimized bioinformatics pipelines. Using 43 samples that were previously validated by Illumina's MiSeq platform and/or by Sanger sequencing/multiplex ligation-dependent probe amplification, we amplified the target with the Oncomine™ BRCA Research Assay and sequenced on Ion Torrent S5 XL (Thermo Fisher Scientific, Waltham, MA, USA). We compared two bioinformatics pipelines for optimal processing of S5 XL sequence data: the Torrent Suite with a plug-in Torrent Variant Caller (Thermo Fisher Scientific), and commercial NextGENe software (Softgenetics, State College, PA, USA). All expected 681 single nucleotide variants, 15 small indels, and three copy number variants were correctly called, except one common variant adjacent to a rare variant on the primer-binding site. The sensitivity, specificity, false positive rate, and accuracy for detection of single nucleotide variant and small indels of S5 XL sequencing were 99.85%, 100%, 0%, and 99.99% for the Torrent Variant Caller and 99.85%, 99.99%, 0.14%, and 99.99% for NextGENe, respectively. The reproducibility of variant calling was 100%, and the precision of variant frequency also showed good performance with coefficients of variation between 0.32 and 5.29%. We obtained highly accurate data through uniform and sufficient coverage depth over all target regions and through optimization of the bioinformatics pipeline. We confirmed that our platform is accurate and practical for diagnostic BRCA1/2 testing in a clinical laboratory.
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1. College of Medicine (의과대학) > Dept. of Laboratory Medicine (진단검사의학교실) > 1. Journal Papers
Yonsei Authors
Kim, Yoon Jung(김윤정) ORCID logo https://orcid.org/0000-0002-4370-4265
Shin, Saeam(신새암) ORCID logo https://orcid.org/0000-0003-1501-3923
Yu, Nae(유내)
Lee, Kyung A(이경아) ORCID logo https://orcid.org/0000-0001-5320-6705
Lee, Seung-Tae(이승태) ORCID logo https://orcid.org/0000-0003-1047-1415
Choi, Jong Rak(최종락) ORCID logo https://orcid.org/0000-0002-0608-2989
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