副猪嗜血杆菌外膜蛋白TbpB基因的克隆和表达

Cloning and expression TbpB gene for outer membrane protein of Haemophilus parasuis

为了克隆和表达副猪嗜血杆菌(Haemophilus parasuis,HPS)外膜蛋白TbpB基因,试验采用基因工程的方法对已发布的HPS外膜蛋白P5序列进行相关序列分析,设计并合成引物,以HPS血清5型基因组为模板,通过PCR扩增HPS外膜蛋白P5编码基因,获得长为1 116 bp的预期基因片段。该基因编码372个氨基酸的蛋白质,预测分子质量约为41 ku,将该基因片段定向克隆至表达载体pGE X-6p-1中。结果表明:诱导表达后分子质量约为67 ku,与副猪嗜血杆菌阳性血清发生了特异性反应。说明该蛋白与抗体发生了反应,有一定免疫原性。

To study the immunogenieity of the proteins related with HPS, A pair of specific primers were designed and synthesized on the basis of the published genome as a template. The TbpB gene of HPS was amplified by PCR, and then was cloned into the expression vector pGEX - 6p - 1 . The fusion protein was successfully expressed, analyzed the immunogenicity of TbpB protein by the Western - blot. The results showed that the TbpB protein was obtained and its molecular weight was about 54 ku. The Western - blot results showed that it had immune response with the HPS positive serum and the protein had immunogenicity. It prepared a basis of subnnit vaccine and diagnostic reagents.