NOVEL USE OF LIMITED PRIMER EXTENSION IN DETECTING MUTATIONS IN HUMANIDURONATE 2-SULFATASE GENE

Hunter syndrome is characterized by a deficiency of iduronate 2-sulfat ase. A large number of mutations in the gene have been reported. We de scribe here the development of a limited primer method for the identif ication of mutations. In the reaction mixture designed for the limited primer extension, one or two deoxynucleotides from the four necessary deoxynucleotides are added as ''selected nucleotides'' and another de oxynucleotide which is radiolabeled is added as the ''limited nucleoti de''. The absence of one or two of the deoxynucleotides limits the len gth of primer elongation, and the low concentration of the ''limited n ucleotide'' causes an ''extension-delay'' effect and results in a band ing pattern upon electrophoresis of the products thus making it possib le to distinguish mutant and normal alleles. We have studied three nov el mutations in exon IX, 407delTT (TTT to T), 423insCC (CCC to CCCCC), and W502X (TGG to TAG) of the iduronate 2-sulfatase gene by the limit ed primer extension method.