鸡致病性大肠杆菌肠毒素(LT)的鉴定及其基因的克隆

Cloning of Heat-labile Enterotoxin Gene of Chicken Pathogenic Escherichia coli

鸡致病性大肠杆菌分离株O78经家兔肠袢结扎试验(RILT)证实,该菌株产生热敏性肠毒素(LT)。用PCR技术从该菌株中扩增出1.2kb的LT基因,然后将纯化的PCR产物克隆到pGEM-T载体中,转化至受体菌JM109中。用Amp/IPTG/X-gal琼脂平板蓝白菌落筛选得到阳性重组菌株,提取质粒用SphI和SalI双酶切鉴定,结果证实,构建的克隆质粒pXCLT1含有LT基因。

Pathogenic avian Escherichia coli O78 is able to produce heat-labile enterotoxin (LT) with rabbit ileumligation test (RILT). Heat-labile enterotoxin gene was amplified from Pathogenic avian Escherichia coli O78 bypolymerase chain reaction (PCR). PCR products were isolated by 1% agarose gel electrophoresis and recovered byPCR preps DNA purification system. Then the heat-labile enterotoxin gene was inserted in pGEM-T vector. Bytransformation of JM109 and screening with Amp/IPTG/X-gal agar plate, we got recombinant strainJM109(pXCLT1). The recombinant plasmid pXCLT1 was studied in detail by restriction endonuclease analysis.Results showed that the recombinant plasmid carried heat-labile enterotoxin gene.