| 傅晶,杨亦荣,倪晓洁,潘晓东,郑建建,郑少玲,林刃舆,蔡明,陈必成.代谢酶基因单核苷酸多态性两对引物-聚合酶链反应技术方法的建立及初步应用[J].Chinese journal of Epidemiology,2009,30(1):63-67 |
| 代谢酶基因单核苷酸多态性两对引物-聚合酶链反应技术方法的建立及初步应用 |
| Establishment and preliminary application of polymerase chain reaction with confronting two-pair primers for the singe nucleotide polymorphisms of metabolic enzymes |
| Received:August 04, 2008 |
| DOI： |
| KeyWord: 单核苷酸多态性 两对引物-聚合酶链反应技术 细胞色素P450酶 环氧化物水解酶 醌氧化还原酶 |
| English Key Word: Polymorphism,singe nucleotide Polymerase chain reaction with confronting two-pair primers Cytochrome P450 Epoxide hydrolase Quinone oxidoreductase |
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| Abstract: |
| 目的 采用相对的两对引物-聚合酶链反应技术(PCR-CTPP),建立简单、准确、快速、经济、大规模检测代谢酶基因单核苷酸多态性(SNPs)的方法.方法 针对I相代谢酶CYP1A1(A4889G)和Ⅱ相代谢酶EPHX1(A416G)、NQO1(C609T)SNPs设计PCR-CTPP引物,优化PCR反应条件,将其基因分型结果与DNA测序结果进行比对,验证准确性.用该PCR-CTPP检测方法对随机选取的183名汉族健康人进行上述SNPs检测,并与其他健康人群进行比较.结果 通过条件优化,PCR-CTPP检测方法可快速清晰地区分CYP1A1(A4889G)、EPHX1(A416G)和NQ01(C609T)的基因型,并与DNA测序结果相符合.183名汉族健康人中,CYP1A1(A4889G):A纯合子103例(56.3%),G纯合子8例(4.4%),A/G杂合子72例(39.3%);EPHX1(A416G):A纯合子142例(77.6%),G纯合子4例(2.2%),A/G杂合子37例(20.2%);NQO1(C609T):C纯合子60例(32.8%),T纯合子32例(17.5%),C/T杂合子91例(49.7%).基因型分布均符合Hardy-Weinberg平衡(P＞0.05),且存在种族差异(P＜0.05).结论 PCR-CTPP检测方法可简单、准确、快速、经济、大规模地检测代谢酶基因SNPs,可用于临床和流行病学大样本筛选. |
| English 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. |
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