人颗粒裂解肽片段在大肠杆菌中的表达  

Expression of Optimised DNA Fragment Encoding Human NKG5 in Escherichia coli

作　　者：李邦印[1] 王臻[1] 李大伟[1] 张灵霞[1] 王治伟[1] 

机构地区：[1]解放军总医院第二附属医院,北京100091

出　　处：《生物技术通讯》2007年第5期750-752,共3页Letters in Biotechnology

基　　金：首都医学发展基金项目(2002-3030)

摘　　要：目的:建立颗粒裂解肽(NKG5)的原核表达载体系统,并在大肠杆菌中获得表达。方法:采用寡核苷酸合成、PCR扩增得到NKG5编码序列,克隆到pGEM-T载体上,经测序正确后,再切下编码序列连接到重组表达载体pGEX-4T-1中,转化大肠杆菌BL21,用IPTG诱导重组工程菌表达,采用谷胱甘肽偶联的Sepharose4B纯化重组蛋白。结果:重组菌株可以表达GST-NKG5融合蛋白,用免疫印迹反应鉴定纯化的融合蛋白,在相对分子质量34000处有一条带。结论:获得了在大肠杆菌中低表达的颗粒裂解肽融合蛋白,为后续研究奠定了基础。Objective： To construct prokaryotic expression vector carrying DNA fragment encoding NKG5 and to be expressed in E.coli. Methods： By PCR, the DNA fragments ecoding NKG5 were obtained and cloned into pGEM-T. DNA fragment sequenced correctly was ligated into pGEX-4T-1. E.coli BL21 strain was transformed by recommended pGEX- 4T-1 and induced by IPTG to express fusion protein GST-NKG5. Results： Expressing plasmid pGEX-4T-1-NKG5 was obtained. The purified fusion protein can reacted to antibody against GST at the protein band whose molecular weight is about 34kD. Conclusion： GST-NKG5 fusion protein is obtained, which will contribute to further research.

关 键 词：颗粒裂解肽 融合蛋白 原核表达 

分 类 号：Q78[生物学—分子生物学]