期刊论文

中文

华中科技大学学报(医学版)

1672-0741

2004

33

2

109-112

10.3870/j.issn.1672-0741.2004.02.001

国家自然科学基金资助项目（No.30300134）;国家自然科学基金资助项目（No.30300134）

本校为其他机构

目的克隆人3种类型载脂蛋白E(apolipoprotein E，apoE)cDNA，表达并纯化3种apoE融合蛋白。方法采用RT-PCR方法从肝组织中克隆得到apoE3 cDNA，以此为基础通过定点突变得到apoE2和apoE4的cDNA。将上述3种cDNA亚克隆到含有His标签的融合表达载体pT7一PL中，表达的蛋白经镍离子柱亲和层析纯化。结果序列分析表明，获得的apoE2、apoE3、apoE4与GenBank中的一致。表达并纯化了N端带有6个组氨酸的apoE2、 apoE3、apoE4融合蛋白。结论 成功地获得了人3种类型apoE cDNA和3种apoE融合蛋白。

Objective To clone three isoforms of apolipoprotein E3 (apoE3) cDNA, to express and purify the corresponding proteins. Methods The cDNA encoding apoE3 was cloned by the method RT PCR from total human liver RNA. Site directed mutagenesis was used to obtain the cDNA encoding apoE2 and apoE4 isforms. These three cDNAs were subcloned into pT7 PL expressing the target protein as a (His)6 tagged fusion. The purified proteins were gained by Ni NTA column. Results These three sequences were confirmed by comparison with GenBank. 6×His fusion proteins (...