酵母双杂交RRS筛选系统中轮状病毒外壳蛋白VP4诱饵载体的构建

发布时间：2018-01-06 18:49

本文关键词：酵母双杂交RRS筛选系统中轮状病毒外壳蛋白VP4诱饵载体的构建　出处：《重庆医科大学》2007年硕士论文　论文类型：学位论文

【摘要】： 酵母双杂交技术自创立以来已成为研究蛋白之间相互作用的重要方法。传统的酵母双杂交所研究的蛋白间相互作用是在细胞核内完成的,而近年来发展起来的酵母双杂交衍生系统---Ras募集系统(RRS),能够将蛋白间相互作用的研究定位在细胞膜上,更具备了灵敏度高、假阳性少等突出优点。在本研究中,我们利用了一对已知蛋白之间的相互作用来验证RRS筛选系统,以期将此筛选系统更广泛地运用于其他蛋白间相互作用的研究以及相互作用蛋白的钓取。轮状病毒(RV)肠炎及其肠道外并发症是威胁儿童健康的重要疾病。RV与其宿主细胞表面受体的结合是病毒感染的第一必备步骤,因此,明确RV受体性质,对该病的预防和治疗有十分重要的意义。现有研究提示,RV外壳蛋白VP4是病毒吸附受体的主要配体蛋白。在本研究中,我们采用RT-PCR法扩增RV外壳蛋白VP4编码基因并将其克隆到Met425-Myc-Ras载体上,构建成适合于RRS筛选系统的诱饵质粒并排除了诱饵的自激活作用,表明我们可以将该诱饵质粒用于RRS筛选系统,为进一步从敏感细胞cDNA文库中钓取RV受体奠定了基础。
[Abstract]:The yeast two hybrid technology since its inception it has become an important method to study on the interaction between proteins. The traditional yeast two hybrid of protein interaction is in the nucleus, and the recent development of the yeast two hybrid system derived ---Ras recruitment system (RRS), to study the interaction between protein localization in the cell membrane, with high sensitivity, low false positive advantages. In this study, we use the interaction between two known proteins to verify RRS screening system, by this screening system more widely used in the research on the interaction between other proteins and proteins interacting with fish take.
Rotavirus enteritis (RV) and parenteral complications are important diseases threatening the health of children with.RV and its host cell surface receptor is the first essential step, so the virus infection, RV receptor, has very important significance for the prevention and treatment of the disease. The present study suggests that RV capsid protein VP4 is the main virus receptor ligand protein adsorption. In this study, we used RT-PCR method to amplify the RV gene encoding the capsid protein VP4 and cloned into the Met425-Myc-Ras vector, to construct the bait plasmid suitable for RRS screening system and exclusion of self activation of bait, that we may put the bait plasmid for RRS screening system. For further from the sensitive cell cDNA library from fishing laid the foundation of the RV receptor.

【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R346

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