三叶因子蛋白在毕赤酵母GS115中高效表达 毕业论文.doc

本科生毕业论文（设计） 题 目：三叶因子蛋白在毕赤酵母GS115 xxx 专业 xxx 年级、班级 2008级 部、系(院) xxxxx 指导教师姓名 专业技术职务 xxx 副教授 2012 年 6 月 4 日 目 录 中文摘要…………………………………………………………………………………1 英文摘要…………………………………………………………………………………2 1引言 ……………………………………………………………………………………3 2材料 ……………………………………………………………………………………4 2.1材料 …………………………………………………………………………………4 2.2主要仪器SDS电泳检测重组蛋白 ……………………………………………………7 3.5筛选高效表达菌株　…………………………………………………………………8 4结果 ……………………………………………………………………………………9 4.1三叶因子蛋白表达菌株的筛选 ……………………………………………………9 4.2三叶因子蛋白的诱导表达 …………………………………………………………9 5讨论……………………………………………………………………………………11 6结论 …………………………………………………………………………………13 参考文献 ………………………………………………………………………………14 附图表 …………………………………………………………………………………15 综述 ……………………………………………………………………………………17 致谢 ……………………………………………………………………………………21  摘 要 目的：GS115中摇瓶培养并甲醇诱导表达，筛 选出三叶因子蛋白高效表达的菌株。 方法：pPICZα为载体，构建三叶因子蛋白真核表达质粒，经SacI 线性化后电转化毕赤酵母GS115感受态细胞，涂布MD平板，得到70个转化子。分别接种到试管中恒温摇动培养24h，再接种到摇瓶BMGY培养基中振荡培养36h，最后转接到BMMY培养基，诱导培养96h，分别在培养24h、48h、72h96h四个时间点SDS电泳检测目的蛋白表达情况，从而筛选出三叶因子蛋白高效表达的菌株GS115诱导表达的最适时间。 结果：SDS电泳，电泳图谱上 结论：GS115获得高效表达。并筛选出高效表 达三叶因子蛋白的工程菌株，为对三叶因子蛋白进一步功能研究奠定了基础。 关键词：GS115；；SDS电泳。 Abstract Objective: Trefoil factor protein in Pichia pastoris yeast strain GS115 shake flasks and methanol induction, screening out high-level expression of trefoil factor protein strains. Methods: To pPICZα trefoil factor protein eukaryotic expression plasmid, the SacI linearized power into Pichia pastoris GS115 competent cells coating the MD plates, 70 transformants. Were inoculated with cultured 24h thermostatic shaking the test tube, and then inoculated into a shake flask BMGY medium in shaking culture 36h, and finally transferred to BMMY medium for induction 96h, respectively, in cultured 24h, 48h, 72h and 96h four Dianbu plus methanol inducible expression of target protein expression by SDSelectrophoresis, which screened the strains of high-level expression of trefoil factor protein. Filter out the engineering strain induced by the above method to culture, explore a trefoil