Pastore L, Caporaso M G, Frisso G, Orsini A, Santoro L, Sacchetti L, Salvatore F
Dipartimento di Biochimica e Biotecnologie Mediche, Facoltà di Medicina e Chirurgia, Università degli Studi di Napoli Federico II, Italy.
Mol Cell Probes. 1996 Apr;10(2):129-37. doi: 10.1006/mcpr.1996.0018.
Duchenne/Becker muscular dystrophy (DMD/BMD) is a severe X-linked myopathy. In 65% of the patients, the mutations responsible for the disease are macrodeletions in the dystrophin-encoding gene that can be identified with multiplex polymerase chain reaction (PCR) technology. We developed a method for quantitative PCR analysis of deletion carriers involving the use of phosphorimager-based scanning of radioactive-labelled PCR products. We calculated the ratios between the areas of two peaks, one corresponding to the deleted segments to be analysed and the other taken as a reference. In carriers, these ratios (R value) were always about half those obtained in normal females. The final diagnostic result, the diagnostic index (DI), is the ratio of the R values between the propositus and a normal subject. We also assessed the variability of each step of the procedure and the overall variability of the DI value, thus obtaining cut-off values that completely discriminated BMD/DMD deletion carriers from normal females. We were also able to classify, as either 'carrier' or 'normal', several females whose status was not identified with linkage analysis.
