Purchase this article with an account.
Shogo Numa, Akio Oishi, Maho Oishi, Tomoko Hasegawa, Kenji Ishihara, manabu miyata, Akitaka Tsujikawa; Step-wise genetic diagnosis for retinitis pigmentosa and Usher syndrome by Next-Generation Sequence. Invest. Ophthalmol. Vis. Sci. 2018;59(9):24.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Retinitis pigmentosa (RP) is genetically heterogenous and more than 80 causative genes are identified to date. Using Next-Generation sequencing (NGS) is now popular to identify causative mutation in such genetically heterogenous disease. However, NGS is still costly and the analysis is labor-consuming. In the present study, we applied a step-wise genetic diagnosis for RP and Usher syndrome consisting of conventional sequencing method and NGS and evaluated the efficiency.
Genetic screening was done in 226 RP or Usher syndrome patients in the following 3 steps; (1) Mutation detection using Taqman probe (for three common mutations in EYS gene; c.8805C>A, c.7919G>A, c.4957dupA and one mutation in RP1L1 gene; c.1972C>T), (2) Whole exome analysis of EYS gene using Illumina Miseq, (3) Whole genome analysis using Illumina Hiseq (In this research, we analyzed all retinal and optical disease genes that had been reported in RetNet at the time of designing this study). Common variants were filtered with allele frequency >0.5% (for recessive variants) or 0.1% (for dominant variants) in healthy cohort. Mutations reported in previous publications or predicted to be loss of function (stop gain, stop loss, frame shift and splice site) are classified as pathogenic. For a novel missense variant, we evaluated the pathogenicity using seven in-silico prediction tools (SIFT,Polyphen2,LRT,MutationTastar,MutationAssesor,GERP++,phyloP). Copy number variants(CNVs), both deletion and duplication, were detected using the read depth-based approach with normalization.
Genetic diagnoses were made in 92/226(40.7%) patients; 10, 31, and 51 patients were diagnosed in step (1), (2), and (3), respectively. Common disease causing genes in the cohort were EYS (40 patients, 17.7%), USH2A (12 patients, 5.3%), RPGR (8 patients, 3.5%), and CNGA1 (5 patients, 2.2%). One patient was diagnosed with CNV analysis; he had one heterozygous deletion CNV and one frameshift single nucleotide mutation in EYS.
Genetic diagnosis for RP and Usher syndrome was made efficiently using step-wise approach.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
This PDF is available to Subscribers Only