人bax真核表达载体的构建及其在人胃癌耐药细胞系SGC7901/VCR中的表达

Construction of human bax eukaryotic expression vector and its expression in human gastric cancer drug resistant SGC7901/VCR cell line

目的：构建人ｂａｘ真核表达载体，经脂质体导入人胃癌耐药细胞ＳＧＣ７９０１／ＶＣＲ，并检测ｂａｘ基因在转导株的表达．方法：采用分子克隆技术将ｂａｘ基因插入真核表达载体ｐＢＫ－ＣＭＶ的多克隆位点之间，以脂质体介导法将目的基因导入受体细胞ＳＧＣ７９０１／ＶＣＲ，Ｇ４１８筛选克隆细胞，Ｗｅｓｔｅｒｎｂｌｏｔ检测ｂａｘ基因的表达．结果：成功地构建了重组质粒ｐＢＫ－ｂａｘ，将之转入细胞后，经Ｇ４１８筛选３０ｄ获得了稳定的细胞克隆，即ＳＧＣ７９０１／ＶＣＲ－ｂａｘｃｅｌｓ．Ｗｅｓｔｅｒｎ印迹证实了ｂａｘ基因转导株具有明显的Ｂａｘ蛋白表达．结论：ｂａｘ真核表达载体的构建，为胃癌耐药的临床治疗打下了基础．

AIM: To construct a human bax eukaryotic expression vector and to detect its expression in human gastric cancer drug resistant SGC7901/VCR cells. HZMETHODS: bax gene was first inserted into polyclonal sites of pBKCMV vector, followed by the transfection of recombinant plasmid into vincristine resistant human gastric cancer SGC7901/VCR cells by lipofectamine. G418 was used to select resistant clones. Then, Western blot was used to detect the expression of Bax protein. RESULTS: Recombinant plasmids were successfully constructed. After they were transfected into human gastric cancer drug resistant SGC7901/VCR cells, stably transfected cell clones were obtained 30 d after G418 selection. Western blot indicated obvious expression of Bax protein in bax transfected cells. CONCLUSION: The establishment of bax gene stable transfectant provides a sound basis for further investigation of gastric cancer multidrug resistance.