维氏气单胞菌鞭毛蛋白flaA的克隆及原核表达  

Cloning and Prokaryotic Expression of Aeromonas veronii Flagellin flaA

作　　者：翟新新[1] 温振才 沈雪飞[1] 孙真[1] 贾生美[1] 张俊辉[1] 王文东[2] 杨振国[1] 卢强[1] 

机构地区：[1]吉林大学人兽共患病研究所,吉林长春130062 [2]吉林大学畜牧兽医学院,吉林长春130062

出　　处：《中国畜牧兽医》2014年第7期40-43,共4页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(30972277)

摘　　要：本试验旨在利用大肠杆菌BL21(DE3)表达维氏气单胞菌鞭毛蛋白flaA,通过免疫印迹反应初步鉴定重组蛋白的抗原性。根据GenBank公布的序列设计1对引物,经PCR扩增获得flaA基因的完整开放阅读框序列,克隆至原核表达载体pET-30a,转入大肠杆菌BL21(DE3)进行诱导、表达和纯化。SDS-PAGE结果显示重组目的蛋白在大肠杆菌成功表达,约在38ku处可见清晰的表达产物,诱导产物大小与理论值相符。经Western blotting检测,结果显示表达的重组蛋白可与特异性血清抗体和His-tag抗体发生免疫反应。因此,本试验成功表达了维氏气单胞菌鞭毛蛋白flaA,为进一步研究其生物学功能及免疫佐剂作用奠定了基础。This study was aimed to express Aeromonas veronii flagellin flaA in E. coli BL21 （DE3） and preliminarily identi-fy antigenicity of recombinant protein through Western blotting. A pair of primers was designed according to the published se-quences in GenBank. Complete ORF sequence of flaA gene was obtained by PCR amplification and cloned into pET-30a vector,and then transformed into E. coli BL21 （DE3） to induce,express and purify. SDS-PAGE results showed that the target proteinwas successfully expressed in E. coli BL21 （DE3） and the product was consistent with the theoretical value and seen clearly atapproximately 38 ku. Western blotting result showed that the expression of recombinant proteins could produce immune reactionwith prepared antibody and His-tag antibody. Therefore,Aerornonas veronii flagellin flaA was successfully expressed, whichlaid the basis for further study the role of biological function and immune adjuvant of flagellin.

关 键 词：维氏气单胞菌 鞭毛蛋白 克隆 表达 

分 类 号：Q786[生物学—分子生物学]