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W. Dailey, K. Drenser, P. Doshi, M. Trese; Development of a Mutation Panel for Screening Mutations in Pediatric Patients With Abnormal Vasculogenisis . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4603.
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© ARVO (1962-2015); The Authors (2016-present)
Abnormal retinal vasculature is present in patients with various pediatric vitreoretinal diseases such as FEVR, Norrie’s, PFVS and ROP. We collected genomic DNA from 160 patients with pediatric retinal diseases(FEVR;n=32, Norries;n=3, ROP;n=96). Three genes (NDP, Fz4 and LRP–5) of the Wnt ß–cantenin signaling pathway have been implicated in vascularization of the retina. A panel of known mutations was selected for these three genes. Appropriate primers were designed and a protocol was optimized to efficiently and accurately screen the selected panel of mutations.
Single base extension technology on a capillary electrophoreses platform was used to screen mutations using genomic DNA. Initially, single mutations were tested individually. PCR and interrogation primers were then designed to construct a multiplex panel. Primer/template concentrations, thermocycling and cleanup conditions were optimized to obtain well separated products. The reactions were separated and analyzed using the Beckman CEQ 8000 genetic analysis system.
The mutation panel was successfully developed. This assay yields reproducible and accurate results.
This Beckman Coulter SNP kit provides and easy and efficient means for screening three genes of the Wnt ß–cantenin signaling pathway.
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