Simultaneous analysis of various mutations on the 21-hydroxylase gene by multi-allele specific amplification and capillary gel electrophoresis.
A detailed study is presented on the detection of various known point mutations using polymerase chain reaction (PCR) based multi-allele specific amplification (MASA) in conjunction with capillary gel electrophoresis (CGE) separation. The resulting PCR products, corresponding to the individual mutations, are labeled with ethidium bromide during CGE separation, and detected by laser-induced fluorescence. MASA proved to be a novel, fast and cost-effective method for simultaneous analysis of multiple known mutation sites, employing more than one allele specific primers in a single PCR reaction. It results in coexisting amplification of numerous DNA fragments differing in size, which are subsequently separated by CGE. In the present study, several point mutations were analyzed simultaneously by MASA-CGE on the 21-hydroxylase gene of a patient with congenital adrenal hyperplasia.
Barta
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Department of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University Medical School, Budapest, Hungary.
Sasvári-Székely
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Guttman
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D000483:Alleles; D010584:Bacteriophage phi X 174; D001483:Base Sequence; D017931:DNA Primers; D019075:Electrophoresis, Capillary / Q000379:methods*; D005456:Fluorescent Dyes; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D015183:Restriction Mapping; D013255:Steroid 21-Hydroxylase / Q000235:genetics*
D017931:DNA Primers; D005456:Fluorescent Dyes; D013255:Steroid 21-Hydroxylase