鸡肺炎克雷伯菌Ⅲ型菌毛A基因序列测定和分析

Sequencing and Analysis of the Type Ⅲ Fimbriae Strucral Gene A of Avian Pathogenic K. pneumoniae

根据GenBank中肺炎克雷伯菌Ⅲ型菌毛结构基因A(mrkA)GI:149187的序列设计一对引物,以5株鸡致病性肺炎克雷伯菌临床分离株的基因组DNA为模板,PCR分别扩增出了543 bp的Ⅲ型菌毛结构基因(mrkA),经纯化、连接、转化、筛选得到含阳性质粒的菌株,将酶切鉴定正确的阳性质粒菌株进行DNA测序并分析。结果表明所扩增的片段为鸡肺炎克雷伯菌mrkA基因,其开放阅读框架为543 bp,编码181个氨基酸。

According to the published sequence of mrkA gene belonging to type 3 fimbriae from GeneBank （GI： 149187） , a pair of primers containing two digestion sites of endolucleases BamH Ⅰ and Xho Ⅰ for was designed and synthesized for mrkA gene amplification. The target gene （mrkA） for encoding type Ⅲ fimbriae with 543 bp in size was amplified by polymers chain reaction （PCR） using the genomic DNA as templates from 5 strains of avian pathogenic K. pneumoniae. Amplifications were cloned into plasmid pMD18-T and the target recombinant plasmids were obtained after purification, ligation, transformation and screening. According to the results of endonuclease digestion and sequencing, the target amplified fragments were mrkA gene from avian K. pneumoniae, which containing 543 bp for encoding 181 amino acid residuals.