RUNX3抑癌基因真核表达载体的构建以及在膀胱癌T24细胞中的表达

Construction of Eukaryotic Expression Vector pIRES2-EGFP-RUNX3 and Its Expression in Human Bladder Cancer T24 Cells

【目的】构建真核表达载体pIRES2-EGFP-RUNX3,并在膀胱癌T24细胞株中表达。【方法】RT-PCR从人正常膀胱组织中获取RUNX3基因并测序,将RUNX3基因克隆进pIRES2-EGFP载体,构建真核表达载体pIRES2-EGFP-RUNX3,并进行EcoRⅠ和XhoⅠ酶切验证。脂质体介导下转染人膀胱癌T24细胞,荧光显微镜检测细胞中增强型绿色荧光蛋白(EGFP)的表达。【结果】成功构建了真核表达载体pIRES2-EGFP-RUNX3,用脂质体转染人膀胱癌T24细胞后,在荧光显微镜下观察到带绿色荧光的人膀胱癌T24细胞。【结论】成功构建RUNX3基因的真核表达载体pIRES2-EGFP-RUNX3并在膀胱癌T24细胞中表达,为进一步研究RUNX3与膀胱癌的关系奠定基础。

[Objective] To construct the eukaryotic expression vector pIRES2-EGFP-RUNX3 and to express it in human bladder cancer T24 cells. [Methods] The RUNX3 gene was amplified by RT-PCR from human normal bladder tissue, and was confirmed by DNA sequencing. The eukaryotic expression vector plRES2-EGFP-RUNX3 was constructed by introducing RUNX3 DNA fragment into the pIRES2-EGFP vector, and was eonfirmed by EcoRI and XhoI digestion analysis. The pIRES2-EGFP- RUNX3 was transfected into human bladder cancer T24 cells using lipofectamineTM 2000. The expressed EGFP was observed under fluorescent microscope. [Results] The eukaryotic expression vector pIRES2-EGFP-RUNX3 was successfully construeted and transfected into human bladder cancer T24 cells. The transfected human bladder cancer displaying green fluorescence was observed under fluorescence microscope. [ Conclusions ] The eukaryotic expression vector plRES2-EGFP-RUNX3 was constructed and EGFP could be expressed in human bladder cancer T24 cells. The present study laid the foundation for the further research of the relation of RUNX3 and human bladder cancer.