PCR-SSCP法分析载脂蛋白E基因多态性

Determining apolipoprotein E gene polymorphism by PCR-SSCP

目的:建立一种快速、简便、敏感的载脂蛋白E(ApoE)基因多态性检测方法。方法:酚/氯仿法提取46例原发性肾病综合征(primarynephroticsyndrome,PNS)患儿全血基因组DNA,聚合酶链式反应(PCR)扩增ApoE第四外显子,取PCR产物10μl加入等体积含有甲酰胺的上样缓冲液中,变性后行8%聚丙烯酰胺凝胶电泳,银染显示结果。同时对PCR产物进行测序,确认单链构象多态性(single strandconformationpolymorphism,SSCP)的检测结果。结果:PCR SSCP法筛选出的各基因型与测序结果完全符合。结论:PCR SSCP法可以快速准确地检测ApoE常见基因型。

Objective ：To establish a fast, convenient and sengtive measure for determining apollpoprotein E （ApoE） gene polymorphism. Methods： The blood genomes in 46 children with primary oephrotic syndrome were extracted by Phenol/Chloroform, the 4th extron of ApoE was amplified by polymerase chain reaction （PCR）, 10 μl PCR products were mixed with the same volume loading buffers containing formylamine, 8% polyacrylamide gel eleetrophoresis after denaturation for silver staining. The sequencings of PCR products were performed to affirm the results of single-strand conformation polymorphism（SSCP）. Results：All genotypes detected by PCR-SSCP were coincident with sequencings perfectly. Conclusion： PCR-SSCP could be quick and accurate means to determine the genotypes of ApoE.