MOLECULAR DIAGNOSIS OF 21-HYDROXYLASE DEFICIENCY - DETECTION OF 4 MUTATIONS ON A SINGLE GEL

Previous studies of the molecular basis of 21-hydroxylase deficiency h ave shown four common gene conversion mutations in exons 7 and 8. Curr ent molecular diagnostic protocols use allele-specific oligonucleotide hybridization (ASOH) to individually detect each of these mutations a nd the corresponding normal alleles. This method is costly, labor inte nsive, and may not provide quantitative results. To expedite molecular diagnosis in families with 21-hydroxylase deficiency, we have designe d and implemented single-strand conformational polymorphism (SSCP) ana lysis. We applied SSCP analysis to 12 families in whom mutations in ex ons 7 or 8 had been previously identified by ASOH. Using a single poly merase chain reaction (PCR) amplification, unique conformers can be as signed to three mutations: V281L, Q318X, and R356W. The fourth mutatio n, T insertion at nucleotide 1761, was detected by heteroduplex analys is of the same PCR product. Thus, we were able to identify all four mu tations using a single PCR product on a single gel. (C) 1994 Academic Press, Inc.