• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

禽多杀性巴氏杆菌外膜蛋白H基因的克隆表达及抗原性鉴定

Cloning, Expression and Antigenicity of Omp H Gene of Pasteurella multocida

  • 摘要: 为了对禽多杀性巴氏杆菌(Pm)外膜蛋白H(Omp H)的免疫学活性进行鉴定,本研究根据已发表的多杀性巴氏杆菌X-73株(U50907.1)设计了两对引物,用PCR方法扩增了禽多杀性巴氏杆菌CVCC44801株的Omp H,扩增片段为1 348 bp(ORF为1 056 bp)。扩增的Omp H与GenBank中登录号为AE004439.1(Pm70株)、EF635422.1(C44-1株)、EU016232.1(P52株)、JQ082510.1(XJ121株)、U50907.1(X-73株)、U52200.1(P-1059株)的序列比对分析结果表明,其核苷酸同源性范围在80.2%~98.4%,氨基酸同源性范围在79.6%~98.5%。扩增了去信号肽的Omp H基因,构建原核重组表达质粒pET-Omp H,转化BL21并诱导表达,SDS-PAGE结果表明,表达蛋白约为56 ku,与预期分子量的大小相符;蛋白免疫印迹试验结果表明,该蛋白具有良好的免疫学活性,为建立禽Pm血清学检测方法以及禽多杀性巴氏杆菌外膜蛋白疫苗的研究奠定了基础。

     

    Abstract: The immunological activity of the outer membrane protein H (Omp H) of Pasteurella multocida (Pm) was identified. Based on the published data on P. multocida strain X-73 (U50907.1), two pairs of primers were used to amplify the Omp H of P. multocida strain CVCC 44801 by PCR.The amplified fragment was determined to be 1 348 bp (ORF at 1 056 bp). The sequence alignment on Omp H of Pm70, C44-1, P52, XJ121, X-73 and P-1059 with those registered in GenBank showed a nucleotide homology between 80.2% and 98.4% and an amino acid homology between 79.6% and 98.5%. The recombinant plasmid pET-Omp H was constructed and transformed into BL21. The expressed protein was about 56 ku, which was close to the expected molecular weight. The western blot also confirmed its immunological activity. The results would provide the information for the establishment of a viable serological detection method and an applicable Omp vaccine against P. multocida.

     

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