Log-PCR: A New Tool for Immediate and Cost-Effective Diagnosis of up to 85% of Dystrophin Gene Mutations

BACKGROUND: Duchenne (DMD) and Becker (BMD) muscular dystrophies are caused by mutations in the dystrophin gene. Despite the progress in the technologies of mutation detection, the disease of one third of patientsescapesmoleculardefinitionbecausethelabor and expense involved has precluded analyzing the entiregene.Noveltechniqueswithhigherdetectionrates, such as multiplex ligation-dependent probe amplification and multiplex amplifiable probe hybridization, have been introduced. METHODS: We approached the challenge of multiplexing by modifying the PCR chemistry. We set up a rapid protocol that analyzes all dystrophin exons and flanking introns (57.5 kb). We grouped exons according to their effect on the reading frame and ran 2 PCR reactionsforDMDmutationsand2reactionsforBMD mutations under the same conditions. The PCR products are evenly spaced logarithmically on the gel (LogPCR) in an order that reproduces their chromosomal locations. This strategy enables both simultaneous mapping of all the mutation borders and distinguishing between DMD and BMD. As a proof of principle, we reexamined samples from 506 patients who had received a DMD or BMD diagnosis.