四环素调控系统对白喉毒素-血管内皮细胞生长因子融合基因表达的影响

Influence of Tetracycline-controlled System on Expression of DT_(390)-VEGF_(165) Fusion Gene

目的：研究四环素调控系统对ＤＴ３９０－ＶＥＧＦ１６５融合基因在人胃癌细胞中表达的影响。方法：构建四环素调控的ＤＴ３９０－ＶＥＧＦ１６５融合基因真核表达载体，用脂质体介导转染胃癌细胞ＳＧＣ７９０１，在四环素有或无的情况下检测胃癌细胞的存活率。结果：４μｇ构建物转染５×１０５个胃癌细胞／孔（２４孔板）９６ｈ后发现，无四环素时转毒素融合基因的细胞的存活率为２０％；四环素存在时转毒素融合基因的细胞的存活率为６６％；转染细胞在四环素有或无的情况下经白喉抗毒素－ＦＩＴＣ免疫荧光抗体染色均呈现黄绿色荧光，表明融合基因在细胞中得到了表达，且四环素调控系统存在基础量的漏表达。结论：四环素调控系统基本上可以控制毒素基因在人胃癌细胞中的表达。

Objective: To study the influence of tetracycline-controlled system on expression of the DT390-VEGF165 fusion gene in human gastric carcinoma cell. Methods: The eukaryotic expression vectors containing DT390-VEGF165 gene under transcriptional c ontrol of a tetracycline-controlled system were constructed. 4μg of constructs were introduced into SGC7901 cells (5×105 cells/well, 24 well) by means of lipofectamine-mediated gene transfer procedure. The transfected cellular morphology was observed bo th in the absence and in the presence of tetracycline. Results: It was shown that virtually 80% SGC7901 cells were killed in the activated state(i.e. without tetracycline), 66% cells were survived in the repressed state(i.e. with tetracycline), whereas a n undesirable leakage of expression occurred in the repressed state. Immunohistochemistry showed the transfected cells with or without tetracycline could be stained by anti-DT-FITC, indicating the chimeric gene expression in SGC7901 cell. Conclusions: Th e chimeric gene can be controlled by tetracycline-controlled system and result in cellular toxicity in the absence of tetracycline.