Cloning of human APOE ε3 gene and construction of its eukaryotic expression vector

GUI Li; et al

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Cloning of human APOE ε3 gene and construction of its eukaryotic expression vector

更新时间：2024-01-17

- Cloning of human APOE ε3 gene and construction of its eukaryotic expression vector
- Journal of Chongqing Medical University Issue 7, Pages: 784-787(2008)
- 作者机构：
  
  1. 泸州医学院附属第一医院眼科
  2. 重庆医科大学附属第一医院神经外科
- 作者简介：
- 基金信息：
- DOI：
  CLC： Q78
- Published：2008，
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[1]郐莉,江涌,孙晓川,郑履平.人载脂蛋白Eε3等位基因的克隆及表达[J].重庆医科大学学报,2008(07):784-787.

GUI Li, et al. Cloning of human APOE ε3 gene and construction of its eukaryotic expression vector. [J]. Journal of Chongqing Medical University (7):784-787(2008)
[1]郐莉,江涌,孙晓川,郑履平.人载脂蛋白Eε3等位基因的克隆及表达[J].重庆医科大学学报,2008(07):784-787. DOI：

GUI Li, et al. Cloning of human APOE ε3 gene and construction of its eukaryotic expression vector. [J]. Journal of Chongqing Medical University (7):784-787(2008) DOI：

摘要

目的:克隆人载脂蛋白E（Apolipoprotein E

APOE）ε3等位基因

测定序列

并构建带有人源性APOEε3等位基因的真核表达载体。方法:通过逆转录聚合酶链式反应（RT-PCR）从人胎儿脑组织中克隆得到APOEε3等位基因

与PMD19-T载体连接

再酶切PMD19-T-APOEε3

获得APOEε3基因

然后将其与表达载体pEGFP-N1连接

构建pEGFP-N1-APOEε3真核表达载体。重组质粒转染293T细胞后

在荧光显微镜下观察EGFP的表达

并用Western-Blot的方法检测目的蛋白在293T细胞中的表达。结果:经酶切和DNA测序证明APOEε3基因序列正确

真核表达载体构建成功。将重组质粒转染293T细胞

在转染后的细胞中检测到目的蛋白的表达。结论:成功构建了pEGFP-N1-APOEε3真核表达载体

该表达载体能在真核细胞中正确表达。

Abstract

Objective:To clone human APOEε3 gene for construction of mammalian expression vector pEGFP-N1-APOEε3. Methods:The cDNA encoding APOEε3 was cloned by the methods RT-PCR from total human embryonic brain RNA. The gene was ligated with PMD19-T vector and then was excised from PMD19-T-APOEε3 plasmid and inserted into pEGFP-N1 to construct eukaryotic expression vector. The expression of APOEε3 gene was identified by western-blot. Results:APOEε3 gene sequence and vector were identified correctly by enzyme digestion and sequence analysis. The APOEε3 gene was transfected into HEK293 cells and expressed successfully. Conclusion:The APOEε3 gene is successfully cloned and its eukaryotic expression vector is obtained

which provides basis for the further experimental researches on APOE functions.

关键词

载脂蛋白E等位基因基因克隆真核表达

Keywords

apolipoprotein EallelesGene cloneMammalian expression

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