可诱导型真核表达载体的构建及其表达效率的测定

目的：利用ＭＴ－Ⅱ启动子的Ｚｎ＾２＋诱导表达特性，构建含多克隆位点的可诱导型真核表达载体ｐＭＤＮＡ３－ｎｅｏ，并用荧光素酶报告基因（ｌｕｃ）检测其表达效率。方法：用分子克隆方法构建载体，并将ｌｕｃ基因克隆到其多克隆位点内，转染胃癌细胞ＳＧＣ７９０１，Ｇ４１８筛选出稳定克隆，分别在不同浓度ＺｎＳＯ４级不同诱导时间下进行诱导表达和测定，并检测不同单细胞株的表达效率。

The construction and the evaluation of expression efficiency of an inducible mammalian expression vector

AIM: To construct an inducible mammalian expression vector pMDNA3neo with multiclone sites using MT promoter, and to analyze the expression efficiency of the vector. METHODS: After the construction of pMDNA3neo by molecular cloning technique, a report gene luciferase (luc) was cloned into its multiclone sites. Stable clones of gastric cancer cell line SGC7901, which have been transfected with pMDNA3luc, were selected with G418, and then the Luc expression was determined at various induction concentrations of ZnSO4 and different time points respectively. The expression efficiency of different monoclone cells was also detected. RESULTS: After inducing at 160 mol/L ZnSO4 for 24 h the Luc expression reached the highest level, which was 27 times higher than the noninduced level (P<0.01). HZCONCLUSION: Mammalian expression vector pMDNA3neo is an ideal vector for induction expression.