| 摘要: |
| 【目的】克隆布鲁氏菌Ⅳ型分泌系统效应分子BPE123和BPE275的基因并进行诱导表达。【方法】从牛种布鲁氏菌2308 株基因组中 PCR 扩增BPE123和BPE275基因片段，亚克隆到 pGEM-T Easy 载体中并测序。将BPE123和BPE275基因克隆至融合表达载体pET-28a，构建表达重组质粒 pET-28a-BPE123和pET-28a-BPE275，在大肠杆菌中进行诱导表达，Western blot分析重组蛋白His-BPE123 和 His-BPE275的免疫学特性。【结果】PCR 扩增获得了462 bp 的BPE123 基因片段和762 bp的BPE275基因片段，成功构建了pET-28a-BPE123、pET-28a-BPE275原核表达载体，并在大肠杆菌中成功表达了BPE123和BPE275蛋白，经 SDS-PAGE 检测，重组融合蛋白BPE123、BPE275 的分子质量分别约为22和31 ku，布鲁氏菌免疫动物血清能特异性识别表达的蛋白。【结论】克隆了分泌蛋白BPE123、BPE275基因片段，并成功表达了布鲁氏菌Ⅳ型分泌系统效应分子 BPE123和BPE275。 |
| 关键词: 布鲁氏菌 Ⅳ型分泌系统 效应蛋白 |
| DOI： |
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| 基金项目:国家自然科学基金项目（31360610）；新疆生产建设兵团国际合作项目（2013BC005）；石河子大学杰出青年项目（2012ZRKXJQ02） |
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| Clone,prokaryotic expression and identification of effector molecules BPE123 and BPE275 of Brucella Ⅳ secretion system |
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SUN Zhihua,ZHANG Hui,LIU Laizhen,et al
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| Abstract: |
| 【Objective】This study aimed to clone and express effector molecules BPE123 and BPE275 of Brucella type Ⅳ secretion system.【Method】The BPE123 and BPE275 genes were obtained from Brucella abortus 2308 genome.Then,they were linked to the vector pGEM T Easy and cloned to expression vector pET-28a to construct recombinant plasmids pET-28a-BPE123 and pET-28a-BPE275.These constructs were expressed on E.coli BL21 cells,and the immunological characteristics were analyzed using Western blot analysis.【Result】Full lengths of BPE123 and BPE275 were 462 bp and 762 bp.Prokaryotic expression vector pET-28a-BPE123 and pET-28a-BPE275 were constructed.The BPE123 and BPE275 protein were successfully expressed in E.coli.SDS-PAGE showed proteins with molecular weights of 22 ku and 31 ku.The expressed proteins can be specifically identified by immune serum against Brucella.【Conclusion】This study successfully cloned BPE123 and BPE275 genes of Brucella abortus 2308 and expressed them. |
| Key words: Brucella type Ⅳ secretion system effector protein |
