K_(99)—F_(41)双纤毛菌的构建

Construction of the Double Fimbriaes Strain for F41 and K99 Two Kinds of Pili Antigenes

产肠毒素大肠杆菌能引起仔猪、犊牛、羔羊等新生幼畜发生急性腹泻、K_(99)和F_(41)是这类产肠毒素大肠杆菌所产生的两种在免疫性质上不同的纤毛抗原。 提取K_(99)质粒,用HindⅢ酶消化,再用低熔点琼脂糖电泳回收分子量约为11Md的K_(99)的HindⅢ片段,并将该片段与经过碱性磷酸酶处理的PBR322载体重组,转化大肠杆菌C_(600)。用ELISA筛选K_(99)抗原阳性克隆。 E.coli83919F_(41)^+是非致病的产生F_(41)纤毛抗原的野生菌株。且F_(41)抗原基因是由染色体编码的。采用转化的方法将K99人工重组质粒转化到E.coli83919F_(41)^+中构建成K_(99)—F_(41)双纤毛基因工程菌。双纤毛菌中K_(99)抗原表达达到给体菌的水平,而F_(41)抗原的表达未受到明显的干扰。

Two immunologically unrelated pili antigen.es, K99 and F41, were produced by enterotoxigenic Escherichia. The strain pathogenisis in piglots, calfes, lambs by its fimbraes and ST enterotoxin. Its fimbriae has immunologition, but do not pathogenisis. Plasmid FTK99-1 was constructed by ligating of 11Md HindⅢ fragment from the K99 plasmid to PBR322 vector. The result of restriction analysis showed that the 11Md fragment from the K99 plasmid was ligated to PBR322 vector. Plasmid PTK99-1 DNA was introduced into E.colic83919 which was the nature strain of expressing F41 antigenes, discoutaining ST enterotoxin, to construct the double fimbriase strain for F41 and K99 two kinds of pili antigenes.