摘要
目的:通过基因工程技术获得重组结核分枝杆茵19 ku蛋白。方法:应用PCR技术扩增卡介苗的19 ku蛋白DNA序列;以质粒pET28a为表达栽体,构建19 ku重组质粒,然后转化大肠埃希菌BL21(DE3);在异丙基硫代-β-D-半乳糖苷(IPTG)诱导下,分别对不同诱导时间的表达产物进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE),凝胶经考马斯亮蓝染色检测蛋白。通过镍柱纯化后获得目的蛋白。结果:重组质粒pET28a-p19测序表明与报道的序列相同。它在大肠埃希菌BL21(DE3)细胞内以可溶性形式表达。不同IPTG诱导时间实验表明重组结核分枝杆茵l9 ku蛋白诱导4 h在大肠埃希菌中的表达量最高。结论:pET28a-p19大肠埃希菌工程株可高表达结核分枝杆菌重组19 ku蛋白。
Objective： To obtain recombinant 19 ku protein from mycobacterium tuberculosis by using gene engineering technology. Methods： Used PCR technology to amplify 19 ku protein DNA sequence of the BCG vaccine; took plasmid pET28a as expression vector, constructed the 19 ku reeombined plasmid, then transfered E.coli BL21 （DE3） , under the inducement of ispropy-β-D-thiogalactoside （IPTG） , took SDS-PAGE to expression product with different induction time, with coomassie brilliant blue dyeing to detect the protein. The recombinant 19 ku protein was purified by Ni-NTA. Results： The sequencing showed the recombinant plasmid pET28a-p19 had the same sequence with reaport. It expressed in soluble form in DE3. The IPTG induction time experiment showed the recombinant mycobacterium tuberculosis 19 ku protein expressed in E.coli reached highest level under 4 hours inducement. Conclusion ： E.coli pET28a-P19 can highly efficiently express the Mycobacterium tuberculosis recombinant 19 ku protein.
出处
《江汉大学学报（自然科学版）》
2012年第6期53-55,71,共4页
Journal of Jianghan University：Natural Science Edition
基金
湖北省教育厅纵向项目(B20093406)
江汉大学大学生科研重点项目
关键词
结核
卡介苗
19
ku脂蛋白
基因表达
tuberculosis
BCG vaccine
19 ku lipoprotein
gene expression
