载脂蛋白E基因型分型方法

Human apolipoprotein ε2/ε3/ε4 genotype determined

目的建立载脂蛋白Ｅ基因型分型聚合酶链反应限制性片段长度多态性方法。方法用特异的探针对氨基酸１１２和１５８多形基因区基因组的ＤＮＡ扩增，产物用ＨｈａⅠ酶消化，其片段在４％琼脂糖凝胶分离。结果经消化的载脂蛋白Ｅ片段在琼脂糖上，ε２／ε２为９１、８３、３８ｂｐ，ε３／ε３为９１、４８、３８ｂｐ，ε４／ε４为７２、４８、３８ｂｐ，ε２／ε３为９１、８３、４８、３８ｂｐ，ε２／ε４为９１、８３、７２、４８、３８ｂｐ，ε３／ε４为９１、７２、４８、３８ｂｐ。３００名献血员其载脂蛋白Ｅ基因分布频率分别为ε２／ε２０％，ε３／ε３７４％，ε４／ε４０％，ε２／ε３１６％，ε２／ε４２％，ε３／ε４８％。结论以聚合酶链反应限制性片段长度多态性方法进行载脂蛋白Ｅ基因型分型，具有清晰、简便。

Objective To establish a polymerase chain reactionrestriction fragment length polymorphism (PCRRFLP) assay of apolipoprotein E (apo E) genotype ε2, ε3 and ε4. Methods Genomic DNA was amplified with specific primers that included the polymorphic region of amino acid 112 and 158. Digestion of the product with Hha I resulted in unique fragments that were separated on agarose.Results The digested apo E fragments on agarose were ε2/ε2: 91, 83, and 38 bp; ε3/ε3: 91, 48 and 38 bp; ε4/ε4: 72, 48 and 38 bp; ε2/ε3: 91, 83, 48, and 38 bp; ε2/ε4: 91, 83, 72, 48 and 38 bp; ε3/ε4: 91, 72, 48 and 38 bp. The relative frequencies of the ε2/ε2, ε3/ε3, ε4/ε4, ε2/ε3, ε2/ε4, ε3/ε4 were 0, 74, 0, 16, 2 and 8 respectively in 300 healthy donors.Conclusions The PCRRFLP assay of human apo E genotype on agarose possesses such advantages as simplicity and rapidity.