Accurate diagnosis of carriers of deletions and duplications in Duchenne/Becker muscular dystrophy by fluorescent dosage analysis.

摘要: We have developed a semiautomated approach to amplify 25 exons of the dystrophin gene using two fluorescent multiplex PCR assays which detect over 98% reported deletions and 90% duplications causing Duchenne/Becker muscular dystrophy. The 5' detects 11 from proximal deletion hotspot while 3' 14 central hotspot. products are accurately sized quantified by DNA sequencer after only 18 cycles amplification. amount product amplified each exon in is divided that other exons, this ratio compared with those control samples obtain series dosage quotients (DQ), copy number determined. No overlap was observed between DQ values obtained single double loci. can be used screen both affected males at risk female relatives for mutation. method has been evaluated as carrier test conducting blind trial on 150 coded samples. Sixty-three carriers, duplication 84 normal controls were all correctly identified, showing diagnosis possible even families where nature mutation unknown. Additionally analysis showed non-pathogenic involving muscle specific promoter 1. Together these 70% mutations gene, greatly simplifying partly automating molecular Duchenne Becker