人口腔细胞P450、GST基因多态性研究  

Use of buccal cells collected in mouthwash as a source of DNA for human genetic polymorphisms analysis

作　　者：程家蓉[1] 关赛芳[1] 王学励[1] 韩丽华[1] 高玉堂[1] 

机构地区：[1]上海市肿瘤研究所,上海200032

出　　处：《肿瘤》2002年第3期242-245,共4页Tumor

摘　　要：目的　探讨人口腔细胞CYP 1A1与 2E1,GSTM1和GSTT1等等位基因的多态性 ,为分子流行病学研究提供一个收集基因样品的简易、可靠的方法。方法　收集人口腔洗液 4 0ml,加 10ml异丙醇固定 ,经蛋白酶K消化 ,酚、氯仿抽提 ,酒精沉淀提取口腔细胞DNA ,结合PCR -RFLP技术分析 32例正常人CYP 1A1与 2E1、GSTM 1和GSTT1基因多态性。结果　32例口腔上皮细胞DNA含量为 2 2 .36～ 330 .70 μg ,均数12 7.38μg ,中位数 113.84 μg。琼脂糖凝胶电泳显示 :所有 32例样品均具有高分子量DNA。其中 31例口腔细胞DNA能扩增 β- globin基因片段。CYP 1A1(Msp1) 3′端侧翼T→C突变型纯合子 (mm)占 2 2 .6 % ,突变型等位基因占 4 8.4 % ,CYP 2E1(Rsa1) 5′端调控区C→T突变型纯合子占 (mm) 6 .4 % ,突变型等位基因占 2 9.0 % ,GSTM1缺失占 4 5 .2 % ,GSTT1缺失 4 8.4 %。结论　从人口腔洗液上皮细胞中可提取到足够量的高分子DNA用以进行多种基因多态性分析 ,是一种可应用于大规模人群进行分子流行病学研究的可靠方法。Objective To develop a simple mouthwash approach to collect buccal cells for obtaining genomic DNA for genetic polymorphism analysis in epidemiological study. Methods Buccal cells were collected by mouthwash of 40ml sterile water, then the mouthwash was added to a 50ml centrifuge tube containing 10ml isopropyl alcohol. DNA was isolated from the buccal cells with a rapid method using proteinase K digestion, phenol-chloroform extraction, and ethanol precipitation. CYP1A1, CYP2E1, GSTM1 and GSTT1 genetic polymorphism were assayed by PCR-RFLP technique. Results The yield of DNA from 32 individual buccal cell collections ranged from 22.36 to 330.70 ug, the average and median was 127.38 and 113.84ug respectively. Electrophoretic analysis showed that the majority of DNA fragments from all 32 samples were in high molecular weight. The β-globin gene fragments were successfully amplified for 31 collections. The 3′- flanking region T to C transition mutant homozygous of CYP1A1 occupied 22.6%, the mutant allele occupied 48.4% ; the 5′- flanking region C to T transition mutant homozygous of CYP2E1 occupied 6.4%, the mutant allele occupied 29.0%. The GSTM1 null occupied 45.2% and GSTT1 null occupied 48.4%. Conclusion The results showed that mouthwash approach is a very useful technique to collect human genomic DNA. The yield of DNA is enough for some clinical and research applications, and the molecular weight is sufficiently high for PCR-based assay. It is particularly suitable for molecular epidemiological study in large population.

关 键 词：口腔洗液 基因多态性 PCR-RFLP CYP1A1 CYP2E1 GSTM1 GSTT1 口腔细胞 分子流行病学 

分 类 号：R78[医药卫生—口腔医学]