RESUMO
Background & objectives: Duchenne (DMD) and Becker muscular dystrophy (BMD) are X-linked recessive disorders, caused by mutations in the dystrophin gene. Genetic diagnosis of the proband becomes crucial, and forms the base for carrier analysis, genetic counselling, prediction of natural history and prognosis, and eligibility for therapeutic strategies. Traditional multiplex PCR assay is the common method used in India to detect DMD gene deletions, mainly in the hot-spot region. Deletions of exons outside the usual 18 or 21 exons in the hot-spot, duplications and carrier analysis are often left without precise genetic diagnosis and require efficient dosage/quantitative analysis. In this study we evaluated the efficacy of using multiplex PCR (mPCR) of 30 exons followed by multiplex ligation-dependent probe amplification (MLPA), to study deletions and duplications in the DMD gene in patients clinically diagnosed as BMD/DMD. Methods: Using an algorithm of mPCR and MLPA which was less invasive and cost-effective, we performed retrospective and prospective analysis on 150 male patients. Results: Multiplex PCR could pick up deletions in 103 of the 150 cases. MLPA was able to detect deletions and duplications including nine additional mutations. Further, the borders of the deletions and duplications were more accurately defined by this recent methodology, which enables one to determine the effect of the mutation on the reading frame. In all, including the single exon deletions, MLPA was efficient in accurately confirming mutations in 35 per cent of all cases. Ten novel mutations were identified in this study. Overall, this approach confirmed mutations in 75 per cent of the patients in our study. Interpretations & conclusions: The systematic approach/algorithm used in this study offers the best possible economical mutation analysis in the Indian scenario.