Integrated DNA, cDNA, and protein studies in Becker muscular dystrophy show high exception to the reading frame rule

Becker muscular dystrophy (BMD) is a milder form of X-linked Duchenne muscular dystrophy (DMD). Here, we report a study of 75 patients with immunoblot and/or immunostaining findings of muscle biopsy consistent with BMD (abnormal dystrophin). We utilized multiplex ligation dependent probe amplification (MLPA) on genomic DNA (gDNA) to screen all 79 exons for both deletions and duplications. A total of 19 patients testing negative for MLPA mutations were tested for mRNA splicing abnormalities using cDNA-MLPA on muscle biopsy. Complete cDNA sequencing was done on MLPA-negative patients. We identified disease-causing mutations in 66 (88%) of the patients. Of the mutation-positive patients, 42 (64%) showed deletions of one or more exons, 14 (21 %) showed duplications, and 10 (15%) showed various mutations detected by cDNA-MLPA and sequencing studies. We found a high rate of "exceptions" to the reading frame rule in this BMD series (out-of-frame BMD; 17/56 deletions/duplications; 30%). This was partly explained by the high incidence of 5' gene deletions in BMD patients (a region known to be a hotspot for exceptions), and due to complex splicing patterns in which a subset of transcripts showed deletions larger than gDNA (exon-skipping). Comparing our findings in BMD to previously published DMD data, BMD patients have higher proportions of duplications, a different distribution of mutations, and higher exception to the reading frame rule.