山西医科大学学报2017，Vol.48Issue(9)：913-917,5.DOI:10.13753/j.issn.1007-6611.2017.09.010
mIDH1真核表达载体的构建及其对U87细胞增殖的影响
Construction of mIDH1 eukaryotic expression vector and its effect on proliferation of U87 cells
段景怡 1王举波2
作者信息
- 1. 宝鸡市中医医院脑病二科,宝鸡721001
- 2. 西安交通大学第二附属医院神经外科
- 折叠
摘要
Abstract
Objective To construct the eukaryotic expression vector of human mIDH1 gene and its stable expression cell line of U87 for investigating its influence on cell proliferation in vivo and in vitro.Methods With cDNA fragment of IDH1 as template,IDH1 fragment was amplified by PCR.Then,IDH1 gene was cloned into the p-EGFP-C1 vector after double digestion and ligation to get wild IDH1 (wIDH1) vector.The wIDH1 vector was used to construct the mutant IDH2(mIDH1) vector using point mutation technology,and the mIDH1 vector was identified by enzyme digestion and DNA sequence analysis.The recombinant plasmids were transfected into U87 cells by lipofectamine 2000.The cell clone that stably expressing mIDH1 was selected out by G418 method and further to get the stable cell line.CCK-8 method was used to analyze the proliferation ability of U87 cell line in vitro,and the tumor formation experiment was used to detect the proliferation of mIDH1 cell line and control U87 cell line in vivo.Results Agarose electrophoresis analysis of plasmid resection and sequencing proved that the recombinant plasmid mIDH1 was constructed successfully.The mIDH1 was transfected successfully and stably expressed in U87 cell line.The mIDH1 inhibited the proliferation of U87 cell line in vitro and slowed down the tumor formation speed in vivo.Conclusion The eukaryotic expression plasmid mIDH1 can be stably expressed in U87 cell line and inhibit the proliferation of U87 cell line in vivo and in vitro.关键词
真核表达载体/胶质瘤/U87细胞/IDH1基因/稳转细胞系/细胞增殖Key words
eukaryotic expression vector/glioma/U87 cells/IDH1 gene/stable expression cell line/proliferation分类
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段景怡,王举波..mIDH1真核表达载体的构建及其对U87细胞增殖的影响[J].山西医科大学学报,2017,48(9):913-917,5.
