肝癌特异性p53基因腺相关病毒载体的构建

目的 构建一种肝癌细胞靶向性的腺相关病毒载体以用于肝癌的促凋亡基因治疗的研究。方法 通过设计含有特定酶切位点的引物，选择地从人基因组中出入甲胎蛋白（α－fe-toprotein,AFP）启动子序列并克隆以真核表达载体pTR－UF5上；再通过平端连接的方法，构建成含甲胎蛋白启动子和人野生型p53基因的重组腺相关病毒（recombinant adeno-associated virus,rAAV）质粒rAAV－AFP－53和绿色荧光蛋白基因GFP（green fluorescence protein）的质粒rAAV－AFP－GFP。结果 成功地构建了以腺相关病毒为载体、受人甲胎蛋白启动子调控表达人野生型p53基因的质粒系统。结论 从理论上说，我们构建的重组腺相关病毒质粒rAAV－AFP－p53，可以特异性地转染到肝癌细胞中从而为肝癌基因治疗提供一种有效手段。

The construction of a hepatoma-specific adeno-associated virus vectorcarrying p53 gene

Objective An adeno associated virus (AAV)vector targeted to hepatoma cells was constructed in order to be used in the apoptosis inducing gene therapy of liver cancer. Methods Firstly, primers containing specific enzyme cutting sites was designed and used to amplify the alpha fetoprotein promoter(AFP promoter) from human genome; Secondly, the promoter was cloned into the eukyrocyte expressing plasmid pTR UF5,resulting in the recombinant AAV vector plasmid containing the reporter gene( rAAV AFP GFP ); thirdly, blunted ligation was applied to construct the recombinant AAV vector plasmid containing the p53 gene( rAAV AFP p53 ). Results Recombinant adeno associated virus plasmid was constructed successfully that carried human wild type p53 gene under the control of human AFP promoter. Conclusion Theoretically the recombinant adeno associated virus plasmid rAAV AFP p53 we constructed could be used in gene therapy for hepatocellular carcinoma.