重组人白细胞介素-2基因痘苗病毒真核表达载体的构建与鉴定  

Construction and Identification of Eukaryotic Expression Vector of Vaccinia Virus Expressing Human Interleukin-2 Gene

作　　者：郜恒骏[1] 朱红音[1] 顾伟齐[1] 楼屹[1] 任卫平[1] 萧树东[1] 

机构地区：[1]上海第二医科大学附属仁济医院,上海市消化疾病研究所200001

出　　处：《胃肠病学》2000年第4期231-233,257,共4页Chinese Journal of Gastroenterology

基　　金：国家自然科学基金!(No.39770345)

摘　　要：目的：构建含人白细胞介素－２（ｈＩＬ－２）基因的痘菌病毒真核表达载体ｐＭＪ６０１，为进一步实施胃癌的基因治疗作必要准备。方法：利用质粒抽提、琼脂糖凝胶电泳、酶切、目的基因与载体的连接、感受态细胞的制备及转化和ＤＮＡ序列分析等多种基因工程技术，将纯化回收的ｈＩＬ－２基因分别与质粒ｐＢｌｕｅｓｃｒｉｐｔⅡＳＫ＋／－及ｐＭＪ６０１进行连接、转化并签定。结果：克隆有ｈＩＬ－２基因的ｐＬＸＳＮ经ＥｃｏＲＩ－ＢａｍＨＩ酶切后，ｈＩＬ－２基因被成功地连接到质粒ｐＢｌｕｅｓｃｒｉｐｔⅡＳＫ＋／－上，再经Ｓａｉｌ－ＢａｍＨＩ酶切后，ｈＩＬ－２基因又被克隆到ｐＭＪ６０１真核表达载体上。结论：重组ｈＩＬ－２基因痘苗病毒真核表达载体ｐＭＪ６０１的构建为胃癌的免疫基因治疗打下了坚实的基础。Background/Aims: To construct eukaryotic expression vector of vaccinia virus, PMJ601, which contains human interleukin-2 (hIL-2) gene for gene therapy of gastric carcinoma. Methods: Genetic engineering techniques such as plasmid extraction, agarose gel electrophoresis, restriction analysis, ligation, preparation of competent cell and transformation, DNA sequence analysis were used to make hIL-2 gene clone to pBluescript Ⅱ SK+/- and pMJ601 and identification. Results: hIL-2 DNA frag- ment from pLXSN with EcoRI-BamH Ⅰ restriction enzymes digest was successfully cloned to pBluescript ⅡSK+/-. In addition, hIL-2 DNA from pBluescript ⅡSK+/- with Sail -BamHI restriction enzymes digest was also successfully cloned to pMJ601. Conclusions: To construct pMJ601 expressing hIL-2 gene is the crucial steps for the basis of gene therapy of gastric carcinoma.

关 键 词：重组人白细胞介素2 痘苗病毒 真核表达载体 胃癌 基因治疗 

分 类 号：R730.5[医药卫生—肿瘤]