Résumé :
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Communication n° 534 Straightforward detectable dystrophin gene rearrangements, such as deletion or duplications involving one entire exon or more, are involved in about 70% of Dystrophinopathies (i.e. Duchenne and Becker muscular dystrophies). In the remaining 30% a variety of point mutations or "small" mutations are suspected. Due to their diversity and to the large size (> 2.7 Mb) and complexity (> 79 exons) of the dystrophin gene, these point mutations are difficult to detect. To overcome this diagnosis issue, we developed an optimized a systematic strategy based on muscle biopsies analyses. Following DNA analysis using QF-PCR (quantitative fluorescent PCR) to detect deletions or duplications, protein extracts from muscle biopsies are analysed by multiplex Western blot, then the full length dystrophin mRNA is amplified by RT-PCR and thouroughly analysed to pinpoint the disease causing abnormalities. Through this approach mutations were identified in all patients for whom the diagnosis of DMD and BMD was suspected and confirmed through disclosing by Western blot of quantitative and/or qualitative dystrophin protein abnormalities. Here we report a total of 94 small mutations that include 34 nonsense, 17 microdeletions, 7 microinsertions, 29 splice mutations, 6 mutations activating cryptic exons and one missense mutation. Mutations are distributed all over the dystrophin gene with an apparent higher frequency (12/80) of mutations in exons 66 and 70, and their flanking splice sites. We discuss the phenotype / genotype correlations with an emphasis on 9 (9/94) nonsense and frameshift mutations associated with BMD or intermediate phenotypes, and report dystrophin mRNA and protein data that support potential molecular mechanisms underlying these unexpected associations. This study highlights the efficacy of the diagnostic strategy herein reported and provide comprehensive phenotype/genotype correlations that take into account consequences of mutations on the expression of the dystrophin mRNA and protein.
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