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 journalArticlepeer-review

56 Scopus citations


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 <20× insufficient coverage were completed by PCR/Sanger sequencing to ensure 100% coverage. We analyzed DNA from 65 unrelated RP patients and detected deleterious mutations in 53 patients with a diagnostic yield of ∼82%.

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
Issue number10
StatePublished - 2014
Externally publishedYes


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

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience


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