细粒棘球蚴原头蚴双向电泳图谱数据库的建立及标记蛋白的定位

发布时间：2018-01-07 21:02

本文关键词：细粒棘球蚴原头蚴双向电泳图谱数据库的建立及标记蛋白的定位　出处：《宁夏医科大学》2011年硕士论文　论文类型：学位论文

【摘要】：目的建立细粒棘球蚴原头蚴双向电泳的图谱数据库;用课题组前期工作得到的细粒棘球蚴原头蚴三种重组抗原制备抗血清,纯化出特异性抗体,以此识别双向电泳图中的蛋白质位点,为后期细粒棘球蚴原头蚴蛋白质组研究中蛋白质的定位、蛋白位置的描述等原头蚴双向电泳图谱的应用奠定基础。方法1样本制备:以细粒棘球绦虫原头蚴为样品;2蛋白分离:应用双向电泳技术对其蛋白质进行分离;3图像采集保存及分析:GS-800扫描仪采集图像,通过bio-rad公司的PDquest软件对得到的原头蚴双向电泳图谱进行分析,初步得出构成原头蚴的蛋白质数量以及蛋白质的分子量和等电点数据;4表达纯化:用细粒棘球蚴原头蚴的三种重组蛋白免疫新西兰家兔制备抗血清,将抗体纯化后进行western blot分析,识别原头蚴双向电泳图谱中的已知蛋白质,利用这些已知蛋白作为图谱中的标记。结果在原头蚴双向电泳图中,分离出240个蛋白质位点,通过软件分析初步得到了每个位点的分子量和等电点信息;成功的纯化出三种细粒棘球蚴原头蚴重组蛋白zw-5,14-3-3,p-29,并采用免疫学的方法制备出抗体,完成了抗体对原头蚴蛋白质组双向电泳图谱的识别。结论1初步建立了细粒棘球蚴原头蚴的双向电泳图谱;2在双向电泳图谱的基础上,通过软件分析得到了240个蛋白质位点,并分析获得了这些蛋白质的分子量和等电点信息;3利用原头蚴重组蛋白制备抗体,在细粒棘球蚴原头蚴的双向电泳图谱上标记了三个已知蛋白靶标。
[Abstract]:Objective to establish a two dimensional electrophoresis map database of Echinococcus granulosus (Echinococcus granulosus). The antiserum was prepared from three recombinant antigens of echinococcus granulosus, and the specific antibody was purified to identify the protein sites in the two-dimensional electrophoretogram. The results laid a foundation for the application of two-dimensional electrophoretic patterns of the proteome of echinococcus granulosus in the study of proteome, such as protein location and protein position description. Methods 1 sample preparation: Echinococcus granulosus was used as sample; (2) protein separation: the protein was separated by two-dimensional electrophoresis. 3Image acquisition, preservation and analysis: GS-800 scanner was used to collect images, and the two-dimensional electrophoresis patterns of protocercaria were analyzed by PDquest software of bio-rad Company. The data of protein quantity, molecular weight and isoelectric point of protein were obtained. 4 expression and purification: three kinds of recombinant proteins of Echinococcus granulosus were used to immunize New Zealand rabbits to prepare antiserum. The antibody was purified and analyzed by western blot. The known proteins in the two-dimensional electrophoretic map of the protocercaria were identified and used as markers in the map. Results 240 protein loci were isolated from the electrophoretogram, and the molecular weight and isoelectric point information of each locus were preliminarily obtained by software analysis. Three recombinant proteins zw-514-3-3p29 were successfully purified and antibody was prepared by immunological method. The antibody was used to recognize the two-dimensional electrophoretogram of protocercaria proteome. Conclusion 1Two-dimensional electrophoretic pattern of echinococcus granulosus was preliminarily established. 2 on the basis of two-dimensional electrophoresis map, 240 protein sites were obtained by software analysis, and the molecular weight and isoelectric point information of these proteins were obtained. 3. Three known protein targets were labeled on the electrophoretogram of E. granulosus by using recombinant protein of E. granulosus.
【学位授予单位】：宁夏医科大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R383

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