Objective: To develop a simple, accurate, rapid, economic, large-scale detection method for the detection of single nucleotide polymorphisms (SNPs) metabolic enzymes, using polymerase chain reaction with confronting two-pair primers (PCR-CTPP).
Methods: The primers of CYP1A1 (A4889G), EPHX1 (A416G) and NQO1 (C609T) were designed for PCR-CTPP, and the PCR conditions were optimized. The results of genotyping were verified by DNA sequencing. The above SNPs were detected by the PCR-CTPP detection method in a randomly selected 183 healthy individuals of Han ethnicity. The genotype frequencies were analyzed and compared with people from other ethnicities.
Results: The allele-specific bands of CYP1A1 (A4889G), EPHX1 (A416G) and NQO1 (C609T) were successfully amplified by PCR-CTPP under the optimal conditions and the results of genotyping were consistent with DNA sequencing. Among 183 healthy Han individuals, the genotypic distributions of CYP1A1 (A4889G) , EPHX1 (A416G) and NQO1 (C609T) showed that the wild-type, homozygous variants, and heterozygotes were 103 (56.3%), 8 (4.4%), 72 (39.3%) and 142 (77.6%), 4 (2.2%), 37(20.2%), 60(32.8%), 32 (17.5%), 91 (49.7%) respectively. The distributions of genotypes were all in accordance with the Hardy-Weinberg equilibrium (P > 0.05), with statistical differences and with other ethnic populations (P < 0.05).
Conclusion: The SNPs of metabolic enzymes can be detected by PCR-CTPP method which is simple, accurate, rapid, economic and with large scale. PCR-CTPP can be used for large scale clinical and epidemiological screening.