Dependable and efficient clinical utility of target capture-based deep sequencing in molecular diagnosis of retinitis pigmentosa

Jing Wang, Victor W. Zhang, Yanming Feng, Xia Tian, Fang Yuan Li, Cavatina Truong, Guoli Wang, Pei-Wen Chiang, Richard A. Lewis, Lee Jun C Wong

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

PURPOSE. The purpose of this study was to establish a fully validated, high-throughput nextgeneration sequencing (NGS) approach for comprehensive, cost-effective, clinical molecular diagnosis of retinitis pigmentosa (RP).

METHODS. Target sequences of a panel of 66 genes known to cause all nonsyndromic and a few syndromic forms of RP were enriched by using custom-designed probe hybridization. A total of 939 coding exons and 20 bp of their flanking intron regions with a total of 202,800 bp of target sequences were captured, followed by massively parallel sequencing (MPS) on the Illumina HiSeq2000 device.

RESULTS. Twelve samples with known mutations were used for test validation. We achieved an average sequence depth of ∼1000× per base. Exons with

CONCLUSIONS. Clinical validation and consistently deep coverage of individual exons allow for the accurate identification of all types of mutations including point mutations, exonic deletions, and large insertions. Our comprehensive MPS approach greatly improves diagnostic acumen for RP in a cost- and time-efficient manner.

Original languageEnglish (US)
Pages (from-to)6213-6223
Number of pages11
JournalInvestigative Ophthalmology and Visual Science
Volume55
Issue number10
DOIs
StatePublished - 2014

Fingerprint

High-Throughput Nucleotide Sequencing
Retinitis Pigmentosa
Exons
Costs and Cost Analysis
Mutation
Point Mutation
Introns
Equipment and Supplies
Genes

Keywords

  • Massively parallel sequencing
  • Next generation sequencing
  • Retinitis pigmentosa
  • Target gene enrichment

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience
  • Medicine(all)

Cite this

Dependable and efficient clinical utility of target capture-based deep sequencing in molecular diagnosis of retinitis pigmentosa. / Wang, Jing; Zhang, Victor W.; Feng, Yanming; Tian, Xia; Li, Fang Yuan; Truong, Cavatina; Wang, Guoli; Chiang, Pei-Wen; Lewis, Richard A.; Wong, Lee Jun C.

In: Investigative Ophthalmology and Visual Science, Vol. 55, No. 10, 2014, p. 6213-6223.

Research output: Contribution to journalArticle

Wang, J, Zhang, VW, Feng, Y, Tian, X, Li, FY, Truong, C, Wang, G, Chiang, P-W, Lewis, RA & Wong, LJC 2014, 'Dependable and efficient clinical utility of target capture-based deep sequencing in molecular diagnosis of retinitis pigmentosa', Investigative Ophthalmology and Visual Science, vol. 55, no. 10, pp. 6213-6223. https://doi.org/10.1167/iovs.14-14936
Wang, Jing ; Zhang, Victor W. ; Feng, Yanming ; Tian, Xia ; Li, Fang Yuan ; Truong, Cavatina ; Wang, Guoli ; Chiang, Pei-Wen ; Lewis, Richard A. ; Wong, Lee Jun C. / Dependable and efficient clinical utility of target capture-based deep sequencing in molecular diagnosis of retinitis pigmentosa. In: Investigative Ophthalmology and Visual Science. 2014 ; Vol. 55, No. 10. pp. 6213-6223.
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AU - Wang, Jing

AU - Zhang, Victor W.

AU - Feng, Yanming

AU - Tian, Xia

AU - Li, Fang Yuan

AU - Truong, Cavatina

AU - Wang, Guoli

AU - Chiang, Pei-Wen

AU - Lewis, Richard A.

AU - Wong, Lee Jun C

PY - 2014

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N2 - PURPOSE. The purpose of this study was to establish a fully validated, high-throughput nextgeneration sequencing (NGS) approach for comprehensive, cost-effective, clinical molecular diagnosis of retinitis pigmentosa (RP).METHODS. Target sequences of a panel of 66 genes known to cause all nonsyndromic and a few syndromic forms of RP were enriched by using custom-designed probe hybridization. A total of 939 coding exons and 20 bp of their flanking intron regions with a total of 202,800 bp of target sequences were captured, followed by massively parallel sequencing (MPS) on the Illumina HiSeq2000 device.RESULTS. Twelve samples with known mutations were used for test validation. We achieved an average sequence depth of ∼1000× per base. Exons with CONCLUSIONS. Clinical validation and consistently deep coverage of individual exons allow for the accurate identification of all types of mutations including point mutations, exonic deletions, and large insertions. Our comprehensive MPS approach greatly improves diagnostic acumen for RP in a cost- and time-efficient manner.

AB - PURPOSE. The purpose of this study was to establish a fully validated, high-throughput nextgeneration sequencing (NGS) approach for comprehensive, cost-effective, clinical molecular diagnosis of retinitis pigmentosa (RP).METHODS. Target sequences of a panel of 66 genes known to cause all nonsyndromic and a few syndromic forms of RP were enriched by using custom-designed probe hybridization. A total of 939 coding exons and 20 bp of their flanking intron regions with a total of 202,800 bp of target sequences were captured, followed by massively parallel sequencing (MPS) on the Illumina HiSeq2000 device.RESULTS. Twelve samples with known mutations were used for test validation. We achieved an average sequence depth of ∼1000× per base. Exons with CONCLUSIONS. Clinical validation and consistently deep coverage of individual exons allow for the accurate identification of all types of mutations including point mutations, exonic deletions, and large insertions. Our comprehensive MPS approach greatly improves diagnostic acumen for RP in a cost- and time-efficient manner.

KW - Massively parallel sequencing

KW - Next generation sequencing

KW - Retinitis pigmentosa

KW - Target gene enrichment

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