江苏大学学报（医学版）2017，Vol.27Issue(6)：466-469,4.DOI:10.13312/j.issn.1671-7783.y170203
含EMC6基因重组真核表达载体的构建及其在人肝L-02细胞中的异位表达
Construction of pcDNA3.1 (+)/EMC6 eukaryotic expression vector and its expression in human liver cell line L-02
Syed Hameed Ali Shah 1何大伟 2何敖林 3李莎莎 3王炳芳3
作者信息
- 1. 江苏大学医学院,江苏镇江212013
- 2. 江苏大学附属昆山医院消化内科,江苏昆山215300
- 3. 江苏大学附属昆山医院临床实验研究中心,江苏昆山215300
- 折叠
摘要
Abstract
Objective:To construct an eukaryotic expression vector pcDNA3.1 (+)/endoplasmic reticulum membrane protein complex subunit 6 (EMC6) and to transfect it into human liver L-02 cell line.Methods:The sequence of EMC6 gene published in Pubmed was analyzed and suitable restriction enzyme cutting site was designed.Human EMC6 gene was amplified by PCR with the cDNA of L-02 cells.The template and the fragment was combined with plasmid pcDNA3.1 (+) by gene recombination technology.After constructing an eukaryotic expression vector pcDNA3.1 (+)/EMC6,the accuracy of construction was verified by colony PCR and sequencing.The recombinant expression vector pcDNA3.1 (+)/EMC6 was transfected into L-02 cells by lipofectamine.The mRNA and protein expression of EMC6 in L-02 cells was respectively detected by RT-PCR and Western blotting.Results:After identification by colony PCR and sequencing,the eukaryotic expression vector pcDNA3.1 (+)/EMC6 was successfully constructed.The RT-PCR and Western blotting results showed the overexpression of EMC6 in L-02 cells.Conclusion:The eukaryotic expression vector pcDNA3.1 (+)/EMC6 was successfully constructed,and can be highly expressed in L-02 cells.关键词
内质网膜蛋白复合体亚基-6/重组质粒/载体构建/转染/L-02细胞Key words
endoplasmic reticulum membrane protein complex subunit 6/recombinant plasmid/vector construction/transfection/L-02 cells分类
医药卫生引用本文复制引用
Syed Hameed Ali Shah,何大伟,何敖林,李莎莎,王炳芳..含EMC6基因重组真核表达载体的构建及其在人肝L-02细胞中的异位表达[J].江苏大学学报（医学版）,2017,27(6):466-469,4.
