L-plastin启动子调控下的肿瘤细胞特异性表达载体的构建及活性分析

Construction and Functional Analysis of a Tumor Cell-Specific Expression Vector Regulated by the L-plastin Promoter

目的:构建肿瘤细胞特异性表达载体pcDNA3.1PLN-GFP及鉴定活性。方法:用2.4kb的L-plastin启动子,以绿色荧光蛋白(GFP)为报告蛋白,通过分子克隆技术,构建真核表达载体pcDNA3.1PLN-GFP,用脂质体介导法转染肿瘤细胞和成纤维细胞后,流式细胞术(FACS)分析载体的肿瘤细胞特异性和活性。结果:仅在表达内源性L-plastin的肿瘤细胞和转化的293细胞中有GFP的表达,发光细胞比率较高,而且L-plastin启动子的活性与CMV启动子活性相当,而在其它细胞中没有检测到GFP的表达。结论:我们构建的pcDNA3.1PLN-GFP是一肿瘤特异性的高效表达载体。

Objective：To construct the tumor cell-specific expression vector pcDNA3.1PLN-GFP and analyze its activity.Methods：The 2.4 kb L-plastin promoter was used as a promoter and green fluorescent protein（GFP）was used as reporter to construct the expression vector pcDNA3.1PLN-GFP with molecular cloning techniques.The recombinant plasmid was transfected into tumor cells and murine fibroblasts with lipofectin.The tumor specificity and activity of this vector were analyzed by fluorescence activated cell sorting.Results：L-plastin GFP protein was expressed only in tumor cells and transformed HEK293 cells that expressed the endogenous L-plastin gene.Its activity was equivalent to that found with a vector containing the CMV promoter.Conclusion：The pcDNA3.1PLN-GFP vector is an efficient tumor cell-specific expression vector.