聚合酶链反应/变性高效液相色谱技术检测载脂蛋白E基因型

Detection of apolipoprotein E genotype by denaturing high performance liquid chromatography and polymerase chain reaction

目的建立快速检测载脂蛋白E(apoE)基因型的变性高效液相色谱技术。方法标准酚-氯仿法提取180份健康人外周血基因组DNA,用聚合酶链反应/变性高效液相色谱(PCR/DHPLC)技术测定apoE基因型,分析人群中apo E基因型和等位基因的分布频率。结果共检出6种基因型:ε3/ε3占73.89%,ε2/ε3和ε3/ε4各占15.56%和7.22%,ε2/ε2、ε2/ε4和ε4/ε4共占3.34%。3种等位基因中,最常见的是ε3,占85.28%,ε2和ε4各占10.00%和4.72%。结论建立了一种基于DHPLC的apo E基因分型方法,该方法具有简便、准确的特点。

Objective To develop a genotyping assay for detection of apolipoprotein E（apo E） genotype by using denaturing high performance liquid chromatography（DHPLC） and polymerase chain reaction（PCR）.Methods Genome DNA samples of 180 healthy subjects were extracted by phenol-chloroform method from peripheral blood.Genotyping assay was developed by PCR/DHPLC to detect the genotypes of apo E gene.Results Among the 6 genotypes determined from the 180 subjects,the most common genotype was ε3/ε3 with frequency of 73.89%,followed by ε2/ε3 and ε3/ε4 with frequency of 15.56% and 7.22% respectively.The total frequency of ε2/ε2,ε2/ε4 and ε4/ε4 was 3.34%.The most common allele was ε3 with frequency of 85.28%,while ε2 and ε4 allele were 10.00% and 4.72% respectively.Conclusion The DHPLC assay developed is simple and accurate for detection of apoE genotype.