欢迎您！东篱公司
退出
-
申报数据库
-
立项数据库
-
成果数据库
-
项目获奖数据库
中文摘要：
目的：构建多房棘球绦虫（Echinococcus muhilocularis，Em）重组双歧杆菌（Bifidobacteria bifidum，Bb）-EmⅡ/3疫苗，并研究EmⅡ/3分子在大肠埃希菌BL21（DE3）中的表达效率。方法：通过PCR扩增EmⅡ/3抗原编码基因；将该基因定向克隆于含有谷胱甘肽-S-转移酶（Glutathione s-transferase，GST）基因的大肠埃希菌-双歧杆菌穿梭表达载体pGEX-1λT，构建重组质粒pGEX—EmⅡ/3；用电穿孔法将该质粒导入Bh，构建多房棘球绦虫重组Bb—EmⅡ/3疫苗。经PCR和酶切鉴定后以IPTG诱导表达EmⅡ／3／GST融合蛋白；SDS—PAGE及Western blot对表达产物进行鉴定。结果：PCR成功扩增出1759bD的EmⅡ/3基因；双酶切证实EmⅡ/3基因插入pGEX-1λT中；PCR和双酶切证实重组Bb—EmⅡ/3疫苗构建成功；SDS—PAGE及Westem blot分析显示重组质粒转化宿主菌在IPTG诱导下高效表达了EmⅡ／3／GST融合蛋白。结论：成功构建了多房棘球绦虫重组Bb—EmⅡ/3疫苗，重组质粒pGEX—EmⅡ/3在大肠杆菌中获得了高效表达，Western blot结果提示表达出的EmⅡ/3重组蛋白具有特异的抗原活性。
英文摘要：
Objective：To construct recombinant Bb-Em Ⅱ /3 vaccine of Echinococcus multilocularis and to investigate the expression efficiency of Em Ⅱ/3 antigen encoding gene in Escherichia coli BL21 （DE3）. Methods：Em Ⅱ/3 antigen gene was amplified by PCR. Then the gene was cloned into Escherichia coli-Bifidobacteria shuttle plasmid pGEX-1 λ T containing gluathione-S-transferaze （GST） gene to construct pGEX-Em Ⅱ /3. The recombinant plasmid was electroporated into Bifidobacteria bifidum（Bb） to construct rBb-EmⅡ/3 vaccine. The vaccine was identified with PCR and restriction-endonuclease digestion. The expression of pGEX-Em Ⅱ/3 was induced with isopropyl-β-D-thiogalactopyranosid （IPTG）,and fusion protein Em Ⅱ/3/GST was examined by SDS-PAGE and Western blot techniques. Results：1 759bp gene of Em Ⅱ/3 was successfully amplified by PCR and cloned into pGEX-1 λ T by restriction analysis, rBb-Em Ⅱ/3 vaccine was successfully constructed according to PCR and restriction-endonuclease digestion.h was demonstrated with SDS-PAGE and Western blot that the fusion protein Em Ⅱ/3/GST were expressed in E.coli,BL21. Conclusions：rBb-Em Ⅱ/3 vaccine of Echinococcus multilocularis is successfully constructed. The plasmid pGEX-Em Ⅱ/3 was highly expressed in E.coli in fused form with GST and this kind of protein shows specific antigenicity according to western blot.
同期刊论文项目
同项目期刊论文
期刊信息
位置：