稳定表达外源野生型p53基因人胆囊癌细胞系的建立和鉴定

目的：建立稳定表达外源野生型p53（wtp53）基因的人胆囊癌细胞系。方法：在脂质体lipofectamine^TM 2000介导下，将含有wtp53的真核表达质粒pCMV-p53导入胆囊癌细胞系GBC—SD中。经G418筛选抗性克隆，扩大培养，建立转染克隆细胞系。用聚合酶链反应（PCR）证实外源野生型p53基因的整合，用反转录聚合酶链反应（RT—PCR）和蛋白印迹法分析目的基因及其蛋白表达。结果：野生型p53基因成功地导入胆囊癌细胞系GBC—SD，转染细胞中存在外源p53基因及蛋白的稳定表达。结论：GBC—SD胆囊癌细胞株能稳定表达外源野生型p53基因。

THE ESTABLISHMENT OF STABLE TRANSFECTION OF HUMAN GALLBLADDER CANCER CELL LINE WITH EXOGENOUS WILD-TYPE p53 GENE

Objective: To investigate exogenous wild-type p53(wtp53) gene transfected human gallbladder cancer cell line.Methods: Eukaryotic expressing plasmid pCMV-p53 containing human wtp53 gene was introduced by lipofectamine-mediated gene transfection into human gallbladder cancer GBC-SD cell lines.After transfection,the cells were selected by G418.Then resistant clones were chosen and expanded in RPMI-1640 culture medium.PCR,RT-PCR and Western blotting were used to determine the presence and expression of exogenous p53 gene.Results: An anti-G418 cell clone was established and expanded in culture.The transfected wtp53 gene GBC-SD-wtp53 cells showed expression of exogenous p53 mRNA and p53 protein.Conclusion: Human gallbladder cancer cell line GBC-SD established in this study expresses consistently exogenous wild-type p53 genes.