J Genet Med.
2008 Jun;5(1):15-20.
Diagnostic testing for Duchenne/Becker Muscular dystrophy using Dual Priming Oligonucleotide (DPO) system
- Affiliations
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- 1Genome Research Center for Birth defects and Genetic Diseases, Asan Institute for Life Sciences, Seoul, Korea. hwyoo@amc.seoul.kr
- 2Medical Genetics Clinic and Laboratory, Asan Medical Center, Seoul, Korea.
- 3Department of Pediatrics, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea.
- 4SeeGene Institute of Life Science, Seoul, Korea.
Abstract
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PURPOSE: Large exon deletions in the DMD gene are found in about 60% of DMD/BMD patients. Multiplex PCR has been employed to detect the deletion mutation, which frequently generates noise PCR products due to the presence of multiple primers in a single reaction as well as the stringency of PCR conditions. This often leads to a false-negative or false-positive result. To address this problematic issue, we introduced the dual primer oligonucleotide (DPO) system. DPO contains two separate priming regions joined by a polydeoxyinosine linker that results in high PCR specificity even under suboptimal PCR conditions.
METHODS
We tested 50 healthy male controls, 50 patients with deletion mutation as deletion-positive patient controls, and 20 patients with no deletions as deletion-negative patient controls using DPO- multiplex PCR. Both the presence and extent of deletion were verified by simplex PCR spanning the promoter region (PM) and 18 exons including exons 3, 4, 6, 8, 12, 13, 17, 19, 43-48, 50-52, and 60 in all 120 controls.
RESULTS
DPO-multiplex PCR showed 100% sensitivity and specificity for the detection a deletion. However, it showed 97.1% sensitivity and 100% specificity for determining the extent of deletions.
CONCLUSION
The DPO-multiplex PCR method is a useful molecular test to detect large deletions of DMD for the diagnosis of patients with DMD/BMD because it is easy to perform, fast, and cost-effective and has excellent sensitivity and specificity.
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