Allele-specific amplification of genomic DNA for detection of deletion mutations: Identification of a French-Canadian tay-sachs mutation

Summary A rapid and efficient method for the detection of a 7.6-kb deletion in the-hexosaminidase A-subunit gene, a mutant allele causing Tay-Sachs disease in French Canadians, is described. The protocol involves PCR (polymerase chain reaction) amplification of target sequences on normal and mutant chromosomes. Three amplification primers, a single 5 primer complementary to normal and mutant DNA templates and two 3 primers specific for normal and mutant DNA templates are required. The primers direct amplification of two unique fragments (normal and mutant) that are easily separated by gel electrophoresis. Allele-specific oligonucleotide hybridization using normal and mutant probes to genomic DNA samples from normal, heterozygous and homozygous individuals confirms these results and is consistent with results of genotypic classification of individuals using Southern analysis. The method is applicable to detection of deletion mutations in cases where some deletion-flanking sequence is known.