Molecular Diagnosis of 21-Hydroxylase Deficiency: Detection of Four Mutations on a Single Gel |
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| Institution: | 1. Department of Civil, Architectural, and Environmental Engineering, Missouri University of Science and Technology, USA;2. School of Civil Engineering and Environmental Science, The University of Oklahoma, USA;3. Department of Mechanical and Aerospace Engineering, Missouri University of Science and Technology, USA;4. Engineering Education Center, Missouri University of Science and Technology, USA;1. Department of Pharmaceutical Chemistry and Technology, Faculty of Pharmaceutical Sciences, Ubon Ratchathani University, Ubon Ratchathani, 34190, Thailand;2. Department of Pharmacognosy, Institute of Pharmaceutical Sciences, University of Graz, Universitaetsplatz 4/1, A-8010, Graz, Austria;3. Department of Pharmacology, College of Pharmacy, Rangsit University, Pathum Thani, 12000, Thailand;4. Department of Electrical and Electronics Engineering, Faculty of Engineering, Ubon Ratchathani University, Ubon Ratchathani, 34190, Thailand |
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| Abstract: | Previous studies of the molecular basis of 21-hydroxylase deficiency have shown four common gene conversion mutations in exons 7 and 8. Current molecular diagnostic protocols use allele-specific oligonucleotide hybridization (ASOH) to individually detect each of these mutations and the corresponding normal alleles. This method is costly, labor intensive, and may not provide quantitative results. To expedite molecular diagnosis in families with 21-hydroxylase deficiency, we have designed and implemented single-strand conformational polymorphism (SSCP) analysis. We applied SSCP analysis to 12 families in whom mutations in exons 7 or 8 had been previously identified by ASOH. Using a single polymerase chain reaction (PCR) amplification, unique conformers can be assigned to three mutations: V281L, Q318X, and R356W. The fourth mutation, T insertion at nucleotide 1761, was detected by heteroduplex analysis of the same PCR product. Thus, we were able to identify all four mutations using a single PCR product on a single gel. |
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