【摘要】 参照GenBank上登录的猪胸膜肺炎放线杆菌血清5 型菌株(AF363361)的apxⅠA基因序列设计了1对特异性引物,用PCR法扩增apxⅠA基因,获得了3 069 bp的片段,将其克隆到pMD 18 T载体,经酶切、PCR鉴定和序列分析,表明克隆是成功的。再将长为1 149 bp的apxⅠA基因的部分片段(编码ApxⅠ的N 端的片段)插入到原核表达载体pET 28(a)中,构建了重组表达质粒pET apxⅠA,转化大肠埃希氏菌JM109(DE3),在IPTG 诱导下获得了高效表达,经SDS PAGE检测,证实表达产物大小约为41 ku。更多还原

【Abstract】 A pair of primers were designed according to the sequence of the apxⅠA gene of Actinobacillus pleuropneumoniae(App, AF363361). The App apxⅠA gene was amplified by PCR. 3069bp of the amplified DNA fragment was cloned into pMD18-T and identified by restriction enzyme analysis, PCR and sequencing. Then 1149bp of DNA fragment of apxⅠA gene encoding N-terminal of apxⅠ was inserted into an expression vector pET-28a to yield recombinant expression plasmid pET-apxⅠA. Finally, the recombinant was transformed into E.coli JM109(DE3) for expression induced with IPTG. The SDS-PAGE analysis showed that the apxⅠA gene was expressed in JM109(DE3) and molecular weight of the expression product was 41ku.更多还原