Biallelic Discrimination Assays for the Three Common Ashkenazi Jewish Mutations and a Common Non-Jewish Mutation,in Tay-Sachs Disease,Using Fluorogenic TaqMan Probes

Abstract

We have developed rapid semiautomated fluorogenic TaqMan assays for the three common Jewish mutations that occur in Tay-Sachs disease, the TATC 4-bp insertion in exon 11 (1,278insTATC), the IVS 12 + 1G → C, splice site mutation in intron 12 (1421 + 1 G → C), and the G → A change at the 3′ end of exon 7 (G269S), as well as for a non-Jewish mutation, IVS9 + 1 G → A, believed to be prevalent in patients of Celtic descent. The TaqMan assays are designed to run on the ABI SDS 7700 sequence detection system, using allele-specific probes that carry a reporter dye at the 5′ end and a quencher dye at the 3′ end. Using a 96-well format, all four assays can be performed simultaneously on the same plate, with real-time fluorescence detection or just an end-point plate read. DNA samples from 78 patients identified as carriers by biochemical screening and genotyped by conventional techniques were used to assess the accuracy and efficiency of the probes in allelic discrimination assays. There were no discrepancies noted between previously assigned genotypes and the results obtained by application of this methodology.