survivin基因启动子驱动的肿瘤特异性腺病毒载体的构建

目的:构建肿瘤特异性启动子驱动的腺病毒载体,检测survivin基因启动子在喉癌细胞中的特异表达活性。方法:PCR扩增survivin启动子,构建分别携带survivin启动子和CMV启动子的真核表达载体pCMV-EGFP和pSurp-EGFP,体外连接法制备重组腺病毒Ad-SP-EGFP和Ad-CMV-EGFP,后转染Hep-2细胞及血管内皮细胞ECV304,荧光显微镜下观察EGFP表达。结果:成功构建survivin基因启动子的腺病毒载体;Ad-SP-EGFP转染后荧光镜下见Hep-2细胞发出较强的绿色荧光,而ECV304细胞无表达。结论:成功制备肿瘤特异性survivin启动子驱动的腺病毒,为进一步开发肿瘤的特异性基因治疗奠定了实验基础。

Construction of a tumor specific adenoviral vector piloted by surviving promoter

Objective:To construct adenoviral vector piloted by tumor specific promoter and to detect the specific expression of survivin gene promoter in laryngocarcinoma cells.Method:The survivin gene promoter was generated by PCR to produce the eukaryotic expression vector pCMV-EGFP and pSurp-EGFP carrying with CMV promoter or survivin promoter.The original recombinant recombinant viruses Ad-SP-EGFP and Ad-CMV-EGFP were harvested by ligation in vitro.The two adenovirus were transfected into Hep-2 cell and vessel endothelial cell ECV304,the expressions of EGFP were detected by fluorescent microscope.Results:Adenoviral vector with survivin gene promoter was successfully constructed.Green fluorescence was observed in Hep-2 cells while not in ECV304 after transfected by Ad-SP-EGFP.Conclusion:Survivin gene promoter piloting adenovirus that we constructed have high specific activity in tumor cells and might be potential in tumor specific gene therapy.