Received on March 31, 2006
Accepted on September 19, 2006

Molecular Diagnostics and Genetics

Quantitative Assay of Deletion or Duplication Genotype by Capillary Electrophoresis System: Application in Prader-Willi Syndrome and Duchenne Muscular Dystrophy

¹ Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan
² Department of Obstetrics and Gynecology and Department of Medical Research, Mackay Memorial Hospital, Taipei, Taiwan
³ Department of Medical Research and Department of Pediatrics, Mackay Memorial Hospital, Taipei, Taiwan
⁴ Department of Medical Genetics and Obstetrics and Gynecology, China Medical University Hospital, Taiching, Taiwan
⁵ Department of Obstetrics and Gynecology, National Taiwan University Hospital, Taipei, Taiwan
⁶ Department of Pediatrics, National Taiwan University Hospital, Taipei, Taiwan
⁷ Institute eGene, Inc., Irvine, CA
⁸ Department of Graduate Institute of Clinical Medicine, College of Medicine, and Department of Medical Genetics, National Taiwan University, Taipei, Taiwan
⁹ Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University Hospital, Taipei, Taiwan

^* To whom correspondence should be addressed. E-mail: ynsu{at}ntumc.org.

Background: Deletions and duplications involving large DNA segments result in underexpression or overexpression, depending on the changes in allele dose, and are known to cause many common disorders. Detection of allele dose variations in the human genome is increasingly important in medical genetic diagnosis.

Methods: We use multiplex quantitative PCR coupled with capillary electrophoresis for accurate allele dose determination. In cases of Prader-Willi syndrome (PWS), a total of 24 patients with PWS, as well as 205 control individuals from the general population, were analyzed by use of multiplex quantitative PCR to amplify the FGFR2 gene, the KRIT1 gene, and the SNRPN gene simultaneously. In cases of Duchenne muscular dystrophy (DMD), we optimized the multiplex quantitative PCR to amplify 38 exons to analyze the DMD gene for rapid diagnosis of 12 DMD-affected males, 12 obligate carriers from families, and 50 unaffected female controls.

Results: We were able to unambiguously diagnose the deletion genotype in PWS patients and identify all deletion or duplication genotypes and carrier status in DMD-affected cases with 100% sensitivity and specificity.

Conclusions: This report describes a novel single assay that can rapidly quantify allele dose to provide accurate clinical genetic diagnosis. This technique offers a valuable alternative for the rapid detection of genomic deletions or duplications and decreases costs because it does not require expensive fluorescent reagents.