Multiplex real-time PCR for detection of deletions and duplications in dystrophin gene
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- 作者：Monica Traverso ; Mauro Malnati ; Carlo Minetti ; Stefano Regis ; Silvana Tedeschi ; Marina Pedemonte ; Claudio Bruno ; Roberto Biassoni and Federico Zara
- 关键词：Duchenne muscular dystrophy ; Carrier diagnosis ; Real-time PCR ; Deletions ; Duplications
- 刊名：Biochemical and Biophysical Research Communications
- 出版年：2006
- 出版时间：6 January 2006
- 年：2006
- 卷：339
- 期：1
- 页码：145-150
- 全文大小：206 K
文摘
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 (ΔΔCt). 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.