title: Multiplex real-time PCR for detection of deletions and duplications in dystrophin gene.
abstract: Genetic testing of Duchenne and Becker muscular dystrophies (DMD/BMD) is a difficult task due to the occurrence of deletions or duplications within dystrophin (DMD) gene that requires dose sensitive tests. We developed three multiplex quantitative real-time PCR assays for dystrophin exon 5, 45, and 51 within two major hotspots of deletion/duplication. Each exon was co-amplified with a reference X-linked gene and the copy number of the target fragment was calculated by comparative threshold cycle method (delta deltaC(t)). We compared the performance of this method with previously described end-point PCR fluorescent analysis (EPFA) by studying 24 subjects carrying DMD deletions or duplications. We showed that Q-PCR is an accurate and sensitive technique for the identification of deletions and duplications in DMD/BMD. Q-PCR is a valuable tool for independent confirmation of EPFA screening, particularly when deletions/duplications of single exons occur or for rapid identification of known mutations in at risk carriers.
authors.affiliation: Unit of Muscular and Neurodegenerative Disease, University of Genova and Institute G.Gaslini, Genoa, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Traverso
authors.forename: Mauro
authors.initials: M
authors.lastname: Malnati
authors.forename: Carlo
authors.initials: C
authors.lastname: Minetti
authors.forename: Stefano
authors.initials: S
authors.lastname: Regis
authors.forename: Silvana
authors.initials: S
authors.lastname: Tedeschi
authors.forename: Marina
authors.initials: M
authors.lastname: Pedemonte
authors.forename: Claudio
authors.initials: C
authors.lastname: Bruno
authors.forename: Roberto
authors.initials: R
authors.lastname: Biassoni
authors.forename: Federico
authors.initials: F
authors.lastname: Zara
mesh_terms: D017931:DNA Primers; D016189:Dystrophin / Q000235:genetics*; D005091:Exons; D005260:Female; D017353:Gene Deletion; D020440:Gene Duplication; D006580:Genetic Carrier Screening; D006801:Humans; D008297:Male; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics*; D016133:Polymerase Chain Reaction
chemical_list: D017931:DNA Primers; D016189:Dystrophin