弗氏2a志贺氏菌2457T株YciD蛋白的融合表达和纯化  

Expression and Purification of the YciD Protein of Shigella flexneri 2a Strain 2457T

作　　者：刘先凯[1] 高美琴[1,2] 孙忠科[1] 冯尔玲[1] 朱力[1] 王恒樑[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071 [2]华中农业大学食品科技学院,湖北武汉430070

出　　处：《生物技术通讯》2009年第1期1-3,共3页Letters in Biotechnology

基　　金：国家自然科学基金(30470101;30700035);国家重点基础研究发展计划(2005CB522904)

摘　　要：目的:原核表达重组弗氏2a志贺氏菌2457T株YciD蛋白,为其功能研究奠定基础。方法:用PCR方法从弗氏2a志贺氏菌2457T株染色体中扩增YciD蛋白编码序列,经过纯化、酶切后克隆到原核表达载体pET32a中,构建重组载体pET32a-yciD,转化大肠杆菌BL21(DE3)菌株获得工程菌株,对其表达和纯化条件进行优化;利用WesternBlot检测融合蛋白的表达。结果:构建了YciD蛋白的融合表达载体,并在大肠杆菌中获得高效表达;经Ni-NTA亲和层析柱纯化获得了高纯度的YciD蛋白;WesternBlot表明,此蛋白可与His标签抗体反应,表明获得了目的蛋白。结论:在原核表达系统中表达、纯化弗氏2a志贺氏菌2457T株YciD蛋白,为进一步对其进行功能研究奠定了基础。Objective： To express and purify YciD protein of Shigella flexneri 2a strain 2457T in vitro. Methods： The gene fragment coding YciD protein was amplified by PCR and inserted into plasmid pET32a to construct a recombinant vector pET32a-yciD, then the recombinant vector was transformed into E.coli BL21（DE3）, and the expression and purification of recombinant YciD protein was optimized. The fusion protein was characterized by Western blot. Results： The pET32a-yeiD vector was successfully constructed and the recombinant YciD protein was purified with Ni-NTA affinity chromatography. Western blot analysis showed that the fusion protein interacted with His-tagged antibody. Conclusion： The recombinant YciD protein was successfully expressed and purified in E.coli. This will be helpful to study the functions of YciD protein in vitro.

关 键 词：弗氏2a志贺氏菌2457T株 YciD蛋白 融合蛋白 表达 纯化 

分 类 号：Q78[生物学—分子生物学]