A QUANTITATIVE POLYMERASE CHAIN-REACTION (PCR) ASSAY COMPLETELY DISCRIMINATES BETWEEN DUCHENNE AND BECKER MUSCULAR-DYSTROPHY DELETION CARRIERS AND NORMAL FEMALES

Duchenne/Becker muscular dystrophy (DMD/BMD) is a severe X-linked myop athy. In 65% of the patients, the mutations responsible for the diseas e are macrodeletions in the dystrophin-encoding gene that can be ident ified with multiplex polymerase chain reaction (PCR) technology. We de veloped a method for quantitative PCR analysis of deletion carriers in volving the use of phosphorimager-based scanning of radioactive-labell ed PCR products. We calculated the ratios between the areas of two pea ks, one corresponding to the deleted segments to be analysed and the o ther taken as a reference. In carriers, these ratios (R value) were al ways about half those obtained in normal females. The final diagnostic result, the diagnostic index (DI), is the ratio of the R values betwe en the propositus and a normal subject. We also assessed the variabili ty of each step of the procedure and the overall variability of the DI value, thus obtaining cut-off values that completely discriminated BM D/DMD deletion carriers from normal females. We were also able to clas sify, as either 'carrier' or 'normal', several females whose status wa s not identified with linkage analysis. (C) 1996 Academic Press Limite d