肿瘤坏死因子α转化酶胞内区诱饵质粒的构建及鉴定  

Construction and Identification of Bait Recombinant Vector of pGBKT7-Tumor Necrosis Factor-α Converting Enzyme Cytoplasmic Tail in the Yeast Two-hybrid System

作　　者：刘鑫[1] 王凤丹[1] 王建醒[1] 赵雪飞[1] 于靖[1] 于晓光[1] 

机构地区：[1]哈尔滨医科大学生物化学与分子生物学教研室,黑龙江哈尔滨150086

出　　处：《中华男科学杂志》2007年第3期229-232,共4页National Journal of Andrology

摘　　要：目的:构建用于酵母双杂交系统的诱饵质粒载体pGBKT7-TACEc(肿瘤坏死因子α转化酶胞内区),并检测其是否具有自身激活及毒性作用。方法:从BALB/c雄性小鼠睾丸组织提取总RNA,用RT-PCR方法获得TA-CEc,克隆于pGBKT7载体上,酶切鉴定及序列分析,并检测pGBKT7-TACEc在酵母双杂交系统中的自激活和毒性作用。结果:实验成功构建了诱饵质粒载体pGBKT7-TACEc,并证明其在酵母双杂交系统中无自激活及毒性作用。结论:pGBKT7-TACEc可应用在酵母双杂交系统中,寻找小鼠睾丸cDNA文库中与TACEc相互作用的蛋白质,为筛选小鼠睾丸中与TACEc相互作用的蛋白奠定了重要基础。Objective ： To construct the bait vector pGBKTT-TACEc （ cytoplasmic tail of tumor necrosis factor-α converting enzyme） of Mathmaker GAL4 Two-hybrid System 3, and to test whether it has self-activation and toxic action. Methods ： TACEc gene was amplified by RT-PCR from the mouse testis, and the EcoRI and BamHI sites were introduced into it. The TACEc gene, after sequenced , was cloned into pGBKTT. Self-activation and toxic action of the recombination vector pGBKTT-TACEc was tested. Results ： The pG- BKTT-TACEc vector was successfully constructed and proved of no self-activation and toxic action. Conclusion： The pGBKTT-TACEc can be applied to the screening of the mouse testis cDNA library in the yeast two-hybrid system.

关 键 词：肿瘤坏死因子α转化酶 睾丸 酵母双杂交系统 自激活作用 

分 类 号：R341[医药卫生—基础医学] Q78[生物学—分子生物学]