| 猪胸膜肺炎放线杆菌外膜蛋白(OMP)5′末端保守区基因的克隆、表达及其免疫活性研究 |
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| 引用本文： | 刘思国,彭永刚,王牟平,王春来,宫强.猪胸膜肺炎放线杆菌外膜蛋白(OMP)5′末端保守区基因的克隆、表达及其免疫活性研究[J].中国预防兽医学报,2004,26(6). |
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| 作者姓名： | 刘思国 彭永刚 王牟平 王春来 宫强 |
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| 作者单位： | 中国农业科学院,哈尔滨兽医研究所,黑龙江,哈尔滨,150001 |
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| 摘 要： | 以猪胸膜肺炎放线杆菌血清7型25-4株基因组DNA为模板,PCR方法扩增外膜蛋白(OMP)5′末端保守区基因片段(OMPc),酶切及核苷酸序列分析鉴定后,与原核表达载体质粒pGEX-6P-1进行连接,构建成重组表达载体pGEX-OM-Pc,转入大肠杆菌BL21中,以IPTG进行诱导,SDS-PAGE电泳分析发现,转化了重组质粒的菌株所表达的融合蛋白相对分子量为34 kD,与实际预测相符,命名为GST-OMPc.GST亲和层析柱进行纯化,ELISA方法对纯化蛋白进行检测.结果表明:纯化蛋白GST-OMPc能够与兔抗猪胸膜肺炎放线杆菌血清7型的阳性血清反应.OMPc蛋白的成功表达为其功能的研究打下基础.
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| 关 键 词： | 猪胸膜肺炎放线杆菌 OMP基因 克隆 表达 |
The cloning,expression and immunogenicity of conservative region fragment of outer membrane protein (OMPc) gene 5′end of actinobacillus pleuropneumoniae |
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| LIU Si_guo,PENG Yong_gang,WANG Mu_ping,WANG Chun_lai,GONG Qiang.The cloning,expression and immunogenicity of conservative region fragment of outer membrane protein (OMPc) gene 5′end of actinobacillus pleuropneumoniae[J].Chinese Journal of Preventive Veterinary Medicine,2004,26(6). |
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| Authors: | LIU Si_guo PENG Yong_gang WANG Mu_ping WANG Chun_lai GONG Qiang |
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| Abstract: | The conservative region fragment of outer membrane protein (OMPc) from actinobacillus pleuropneumoniae (APP) 25_4 strain chromosomal DNA was amplified by PCR technique.PCR product was cloned into pGEX_6P_1 expressing vector.Plasmid DNA (pGEX_OMPc)was extracted and digested with enzymes and sequenced to confirm its rightness.The right recmbinant plasmids pGEX_OMPc were transformed into E.Coli BL21 strain.Bacterial lysates prepared from 1?mmol/L IPTG induced cultures were loaded directly for SDS_PAGE.The result shows the recombinant pGEX_OMPc produced a fusion protein (GST_OMPc) with an apparent MW of 36 KDa.The fusion protein reacted with rabbit anti_pig positive sera in ELISA.In conclusion,we obtained the recombinant pGEX_OMPc expressing vector which contains OMPc gene and can be expressed successfully. |
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| Keywords: | Actinobacillus pleuropneumoniae OMP gene cloning expression |
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