大肠杆菌F18ac菌毛F亚单位基因的克隆与序列分析

Cloning and Identification of the Minor Subunit fedF Gene of the Fimbriae F18ac of E.Coli

根据已经发表的 F1 8ab菌毛 F亚单位(Fed F/ ab)的基因 (fed F/ ab) ,设计一对引物 ,利用 PCR技术从表达 F1 8ac菌毛的大肠杆菌2 1 3 4P株、81 99株、881 3株中分别扩增到一段序列 ,并克隆至 p GEM-T载体 ,获得重组质粒T881 3 F、T81 99F、T2 1 3 4PF。琼脂糖凝胶电泳、序列测定及分析表明 ,该 3个序列大小均为 90 3bp,与 fed F/ ab大小一致且具有较高的同源性(99.4% ) ,推导的 Fed F/ ac氨基酸序列与 Fed F/ab同源性为 98.3 %。数据表明该实验所克隆的序列均为 F1 8ac菌毛 F亚单位 (Fed F/ ac)的基因 (fed F/ ac)。

A pair of primers were designed and synthesized according to fedF of fimbriae F18ab (fedF/ab), and the coordinate subunit genes of F18ac (fedF/ac) were amplificated from three different Escherichia coli strains, 2134P?8199 and 8813, which are all display fimbriae F18ac. The PCR products of fedF/ac were cloned into pGEM-T vector. Color screening, PCR and restriction endonucleases analysis were used to identify the recombinant plasmids, and three recombinant plasmids, T8813F, T8199F and T2134PF were obtained. The three recombinant plasmids sequences were analyzed (with the MicroGenic computer program), and compared with the published sequences of fedF/ab. The data of sequences in the three fedF/ac genes showed that their sizes were in accord with the sequences of fedF/ab, and all shared 99.4% and 98.3% identity at nucleoide and amio acid level,respectively. All the data in this study conformed the three genes derived from the three different Escherichia coli strains, 2134P, 8199 and 8813, are all fedF/ac genes of F18ac.