结核分枝杆菌16kDa与GST融合表达及纯化  

Construction and Preparation of Mycobacterium tuberculosis 16kDa Protein

作　　者：郭庆水[1] 朱中元[2] 罗越华[1] 王海波[2] 黄惜[1] 刘贤杰[1] 

机构地区：[1]海南大学农学院,海南海口570228 [2]海南医学院附属新华医院,海南海口570311

出　　处：《生物技术》2009年第1期17-19,共3页Biotechnology

基　　金：海南省重点科技项目("结核抗体检测蛋白芯片系统的研制";080209)资助

摘　　要：目的:构建融合表达GST和结核分枝杆菌16kDa蛋白的表达系统,摸索表达条件及制备方法。方法:根据16kDa蛋白的基因序列设计引物,从结核菌基因组DNA中扩增该基因,酶切后插入表达载体,测序证实后转入大肠杆菌JM109菌株,经IPTG诱导表达,产物经谷胱甘肽-琼脂糖凝胶法纯化。结果:16kDa基因产物大小、酶切片断大小和载体的大小与设计一致、16kDa基因测序的结果与目的基因完全符合。表达纯化的蛋白的大小与蛋白质分子量标准相比较一致。结论:构建的表达载体成功,纯化的蛋白即为目的蛋白。Objective：To construct the expression system for 16kDa of M. tuberctdosis and establish its methods for expression and purification. Method：Primers containing endoneaclease cleavage sites were designed according to M. tubrculosis 16kDa gene sequence and to amplify the16kDa gene. After endonuclease digestion, 16kDa gene was inserted into the expression vector of p - GEX - 6P- 1 to construct 16kDa expressing vector 16K- pG. 16kDa was prepared with IPTG induction and analyzed by SDS- PAGE. Various conditions were tested for maximal production of 16kDa. Result：The vector 16K- pG was comfirmed by endonuclease digestion and DNA sequencing. The 16kDa was demonstrated by SDS - PAGE. It was found that maximal preparation of 16kDa could be obtained at 37℃ with 0.5μmol/mL IPTG for culturing 3h. Pure target protein has been gained with the protocol we developed. Condusions： 16K - pG was proved to express 16kDa of M tubeculosis sucessfully.

关 键 词：结核分枝杆菌 16kDa 表达 纯化 

分 类 号：Q786[生物学—分子生物学] Q785