猪胸膜肺炎放线杆菌整合膜蛋白基因的克隆与表达

Cloning and expression of an integral membrane protein gene of Actinobacillus pleuropneumoniae

为了研究猪胸膜肺炎放线杆菌(APP)的保护性免疫机制,试验采用筛选APP3～8kb随机基因组文库的方法获得1株阳性克隆,经测序拯救质粒得到序列,通过Blast分析确定此片段为APP整合膜蛋白(IM)基因,预计成熟蛋白分子质量为48.431ku。根据序列设计特异性引物并PCR扩增该基因,分子克隆表达该整合膜蛋白。Western-blot分析表明,该蛋白与APP康复期血清发生反应,说明该蛋白在免疫应答中起作用。

To study the immune protective mechanism ofActinobacillus pleuropneumoniae（ APP）, an integral membrane protein （IM） was identified by screening a phage library of 3 -8 kb random DNA fragments of APP. The inserted segment encoding the entire amino acid sequence of the IM was determined by sequencing the rescued plasmids and Blast analysis. Specific primers were designed to amplify the IM gene. The IM was cloned, expressed and purified. Western -blot result showed strong reactivity of the protein with convalescent sera （IgG）. These suggest IM plays a role in immune response.