熔解曲线分析法检测乙醛脱氢酶2基因多态性

Detecting acetaldehyde dehydrogenase2 gene polymorphisms by a melting curve method

目的应用熔解曲线分析法建立乙醛脱氢酶2(ALDH2)基因Glu504Lys多态性分型方法并探讨其临床应用价值。方法提取受检者EDTA-K2抗凝外周血的基因组DNA,用等位基因特异性PCR(AS-PCR)扩增ALDH2基因,通过熔解曲线分析扩增峰,对乙醛脱氢酶2基因504位点的多态性进行基因分型,并与DNA微阵列芯片法的检测结果进行比较,出现结果不一致时,用基因测序法进行验证。结果对60例外周血标本进行ALDH2基因的多态性分型检测,两种方法检测结果均为野生型(Glu504Glu)27例,突变型(Lys504Lys)13例,杂合型(Glu504Lys)20例,熔解曲线分析方法与DNA微阵列芯片法分析结果完全一致(Kappa值为1)。结论本研究成功建立了基于熔解曲线分析检测乙醛脱氢酶2基因Glu504Lys多态性分型方法。

Objective To develop a melting curve method for the genotyping of the acetaldehyde dehydrogenase 2（ALDH2）Glu504 Lys genotype and discuss its clinical application value. Methods The genomic DNA was extracted from peripheralblood cells,and ALDH2 gene was amplified with allele specific PCR（AS-PCR）. The genetype of ALDH2 polymorphismwas determined by amplification peak melting curve,and compared with the by gene chip method. DNA sequencing methodwas used to verify the genotype when the results of two methods were different. Results The results of melting curvemethod were completely matched with the results of gene chip method. Among 60 samples,27 were wild type（Glu504Glu）,13 were hybrid type（Lys504Lys）,and 20 were mutant type（Glu504Lys）. Results of Melting curve analysis and the genechip analysis were identical（Kappa =1）. Conclusion Method based on the melting curve analysis was able to determinethe polymorphisms of ALDH2 gene Glu504Lys genotype.