Abstract
CYP2C9 catalyzes the metabolism of important drugs such as phenytoin, S- warfarin, tolbutamide, losartan, and nonsteroidal anti-inflammatory drugs. A functional polymorphism of the CYP2C9 gene has been described. The single- base mutation of A1061C (Ile359Leu) in the CYP2C9 gene termed CYP2C9*3 was found at a frequency of about 2.1% in Japanese. We developed a rapid mutation analysis method for detecting the CYP2C9*1 genotype. This method is a marriage of two emerging technologies: allele-specific amplification primers for target DNA and a new double-stranded DNA-selective fluorescent dye, SYBR Green. Genotypes are separated according to the different threshold cycles of the wild-type and mutant primers. We applied this procedure to DNA extracted from the blood of healthy Japanese volunteers. The CYP2C9 wild-type CYP2C9*1/CYP2C9*1 and heterozygous CYP2C9*1/CYP2C9*3 genotypes of the CYP2C9 alleles detected by the assay were consistent with the results obtained from restriction enzyme cleavage. No genotype of CYP2C9*3/CYP2C9*3 was found in these samples. Using plasmid DNA containing a point mutation of CYP2C9*3 as template, the assay separated the three genotypes. We conclude that this simple, rapid, and inexpensive procedure is applicable to routine high-throughput assays.
Original language | English |
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Pages (from-to) | 357-362 |
Number of pages | 6 |
Journal | Molecular Genetics and Metabolism |
Volume | 68 |
Issue number | 3 |
DOIs | |
Publication status | Published - 1999 Nov |
Keywords
- Allele-specific amplification
- CYP2C9
- Genetic polymorphism
- P450
- PCR
- SYBR Green
ASJC Scopus subject areas
- Endocrinology, Diabetes and Metabolism
- Biochemistry
- Molecular Biology
- Genetics
- Endocrinology