出血性大肠杆菌O157:H7的Tir与Hly融合基因的构建和表达  

Construction and expression of fusion protein Tir-Hly from enterohemorrhagic Escherichia coli O157:H7

作　　者：张雪寒[1] 何孔旺[1] 卢维彩[1] 李丽[1] 茅爱华[1] 周萍[1] 

机构地区：[1]江苏省农业科学院兽医研究所,江苏南京210014

出　　处：《中国兽医学报》2010年第5期647-650,共4页Chinese Journal of Veterinary Science

基　　金：国家支撑计划基金资助项目(2007BAD40B01)

摘　　要：构建表达Tir和Hly的融合基因,将Tir基因的C端414个氨基酸残基(Tir414)基因部分与Hly基因的C端300个氨基酸残基(Hly300)基因部分串联构建pET28a-Tir414-Hly300重组质粒,将其转化于BL21(DE3),用IPTG进行诱导表达,经SDS-PAGE电泳检测,该融合蛋白获得了高效表达。薄层扫描分析表明,目的蛋白表达量占菌体总蛋白含量的30%左右。由于该融合蛋白由Tir和Hly2部分抗原组成,可刺激机体产生针对转位紧密素受体(Tir)和溶血素(Hly)的抗体,在EHECO157亚单位疫苗设计或单克隆抗体抗制备中具有重要价值。To construct tir-hly fusion gene,the DNA fragments encoding C terminal 414 amino acids（Tir414）of tir and C terminal 300 amino acids（Hly300）of hly were fused in turn in frame with the multiple cloning sites of pET-28a,and the recombinant plasmid of pET28a-tir-hly was transferred to host cells E.coli BL21 strain （DE3）.Then,the protein was induced and expressed with IPTG.The expression quantities and style of fusion protein were determined by SDS-PAGE and thin layer scanning results showed that amount of expressed fusion protein was 30% in total of bacteria protein.Because the fusion protein was consisted of tir414 and hly300 antigens,it can evoke immune responses against tir414 and hly300.The tir-hly fusion protein expressed in E.coli could offer an important ingredient that should be consider in construction of recombinant EHEC O1 57 vaccine or preparation of experimental material for the monoclonal antibody.

关 键 词：EHECO157:H7 转位紧密素受体 溶血素 融合基因 

分 类 号：S852.61[农业科学—基础兽医学]