酸弗林蛋白酶酸性氨基酸簇分选蛋白-1的原核表达、纯化及多克隆抗体制备  

Prokaryotic Expression and Purification of Phosphofurin Acidic Cluster Sorting Protein 1 and Preparation of its Polyclonal Antibody

作　　者：李英辉[1] 刘忠湘[1] 赵亚[1] 黄豫晓[1] 刘军[1] 薛采芳[1] 万磊[1] 

机构地区：[1]第四军医大学病原生物学教研室,陕西西安710032

出　　处：《生物技术通讯》2008年第3期362-364,共3页Letters in Biotechnology

基　　金：国家自然科学基金项目(39970038)

摘　　要：目的:原核表达和纯化PACS-1,并制备其多克隆抗体。方法:通过RT-PCR扩增出PACS-1的编码基因,测序正确后克隆入原核表达载体pGEX4T-1,转化大肠杆菌BL21(DE3),以IPTG诱导PACS-1与GST融合蛋白的表达并经Glu-tathione Sepharose 4B纯化;经SDS-PAGE和Western blot鉴定,应用纯化的蛋白免疫家兔制备多克隆抗体,用ELISA测定抗体的效价。结果:表达和纯化了PACS-1,并获得了较高效价的抗血清。结论:获得纯化的PACS-1及其多克隆抗体,为进一步研究PACS-1的功能奠定了基础。Objective： To express and purify phosphofurin acidic cluster sorting protein 1 （PACS-1） in E.coli and to prepare rabbit anti-pACS-1 antibody. Methods： PACS-1 gene was amplified by RT-PCR and cloned into prokaryotic expression vector pGEX4T-1. The recombinant vector was transformed into E.coli BI221（DE3）. PACS-1 expression was then induced by IPTG. The expressed GST-PACS-1 fusion protein was purified by Glutathione Sepharose 4B and characterized by SDS-PAGE and Western blot. The purified protein was injected into a rabbit, and the titer of the rabbit＇s anti-serum was measured by ELISA. Results： The PACS-1 was successfully purified and the rabbit＇s anti-serum against PACS-1 with high titer was obtained. Conclusion： The preparation of the purified PACS-1 and its polyclonal antibody can be used for further investigation.

关 键 词：PACS-1 原核表达 多克隆抗体 

分 类 号：Q78[生物学—分子生物学]