Establishment and preliminary application of polymerase chain reaction with confronting two-pair primers for the singe nucleotide polymorphisms of metabolic enzymes / 中华流行病学杂志

ABSTRACT

Objective To develop a Simple,accurate,rapid,economic,large-scale detection method for the detection of singe 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.