绵羊肺炎支原体多表位融合基因MO-meAg1的构建、表达及重组蛋白免疫原性分析

发布时间：2018-11-20 18:24

【摘要】：为获得具有良好免疫原性的绵羊肺炎支原体(MO)重组蛋白,利用ABCpred等软件预测分析MO EF-P、PDHA、P97相似蛋白和EF-Ts蛋白的优势线性抗原表位,构建多表位融合基因MO-meAg1,合成后将其亚克隆至表达载体pET-32a(+),构建原核表达载体pET-32a(+)-MO-meAg1。将pET-32a(+)-MOmeAg1转化至感受态细胞E.coli BL21(DE3),IPTG诱导表达后检测重组蛋白反应原性和免疫原性。SDSPAGE结果显示,MO-meAg1重组蛋白分子质量约34.5ku;Western blot分析结果显示,该重组蛋白可被MO阳性血清特异性识别,具有良好的反应原性。小鼠免疫试验结果表明,重组蛋白MO-meAg1具有较强的免疫原性,诱导机体产生的抗血清其间接血凝效价可达1∶128以上。获得的多表位融合蛋白MO-meAg1为绵羊支原体肺炎的免疫学诊断及新型疫苗研究奠定基础。
[Abstract]:In order to obtain (MO) recombinant protein of Mycoplasma pneumoniae of sheep with good immunogenicity, the dominant linear epitopes of MO EF-P,PDHA,P97 similar protein and EF-Ts protein were predicted and analyzed by ABCpred and other software. Construction of multiple epitope fusion gene MO-meAg1, synthesis and subcloning into expression vector pET-32a (), to construct prokaryotic expression vector pET-32a ()-MO-meAg1. PET-32a ()-MOmeAg1 was transformed into E.coli BL21 (DE3), IPTG induced expression). The results of SDSPAGE showed that the molecular weight of MO-meAg1 recombinant protein was about 34.5 ku. The results of Western blot analysis showed that the recombinant protein could be specifically recognized by MO positive serum and had good reactivity. The results of mouse immunological test showed that the recombinant protein MO-meAg1 had strong immunogenicity, and the indirect hemagglutination titer of the antiserum induced by the recombinant protein was over 1: 128. The obtained multiepitope fusion protein MO-meAg1 lays the foundation for the immunological diagnosis of Mycoplasma Sheep pneumonia and the study of novel vaccine.
【作者单位】：石河子大学动物科技学院;中国农业科学院兰州兽医研究所;
【基金】：国家国际科技合作专项(2014DFR31310) 国家自然科学基金(31360596,30960274)~~
【分类号】：S852.62

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