HBV靶向核糖核酸酶基因的原核表达和纯化及多克隆抗体的制备  

Prokaryotic expression and purification of HBV targeted ribonuclease gene and preparation of its polyclonal antibody

作　　者：李英辉[1] 刘军[1] 赵亚[1] 薛采芳[1] 丁劲[1] 黄豫晓[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：《医学研究生学报》2005年第4期295-297,301,共4页Journal of Medical Postgraduates

基　　金：国家自然科学基金资助项目(批准号:30100157)

摘　　要：目的:旨在获得HBV靶向核糖核酸酶,并制备其兔的多克隆抗体。方法:将构建好的HBV靶向核糖核酸酶基因克隆入原核表达载体pET32a(+),转化入大肠杆菌BL21(DE3),以IPTG诱导6×His融合蛋白的表达并经Ni2+亲和层析纯化。通过SDS PAGE和Westernblot鉴定后,用纯化的蛋白免疫家兔,制备多克隆抗体,并以复性的蛋白作为抗原,用ELISA测定抗体的效价。结果:成功地纯化和复性了HBV靶向核糖核酸酶,获得了较高效价的抗血清。结论:获得纯化并复性的HBV靶向核糖核酸酶和多克隆抗体,为进一步研究奠定了基础。Objective: To express and purify HBV targeted ribonuclease (TR) gene in E.coli and to prepare rabbit anti-HBV TR antibody. Methods: HBV TR gene was inserted into prokaryotic expression vector pET32a(+). The recombinant vector was transformed into BL21(DE3). HBV TR expression was then induced by IPTG. The expressed His-tagged protein was purified by Ni 2+ affinity chromatography column 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, using the renatured HBV TR as antigen. Results: The HBV TR was successfully purified and refolded. And the rabbit's anti-serum against HBV TR with high titer was obtained. Conclusion: The preparation of the purified and refolded HBV TR and polyclonal antibody can be used for further investigation.

关 键 词：HBV靶向核糖核酸酶 原核表达 多克隆抗体 

分 类 号：R373.21[医药卫生—病原生物学]