| 副猪嗜血杆菌ompP2基因的克隆、表达及间接ELISA抗体检测方法的建立 |
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| 引用本文： | 贾爱卿,李春玲,王贵平,何永龙,袁子彦.副猪嗜血杆菌ompP2基因的克隆、表达及间接ELISA抗体检测方法的建立[J].中国兽医学报,2011,31(9). |
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| 作者姓名： | 贾爱卿 李春玲 王贵平 何永龙 袁子彦 |
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| 作者单位： | 广东省农业科学院兽医研究所,广东广州,510640 |
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| 基金项目： | 广州市科技计划项目(2010Y1-C311) |
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| 摘 要： | 用PCR方法扩增副猪嗜血杆菌的外膜蛋白ompP2基因,序列测定结果表明扩增片段全长1 188bp。将扩增片段插入表达载体pET-32a,转化BL21进行诱导表达,SDS-PAGE电泳结果证实重组融合蛋白约为60 000,West-ern-blot结果表明重组表达蛋白具有免疫反应性。以纯化的重组蛋白ompP2作为抗原,建立副猪嗜血杆菌的抗体间接ELISA检测方法。通过试验确定最佳反应条件为抗原包被质量浓度4.75mg/L,4℃包被过夜,待检血清的稀释浓度为1∶40,阳性判断标准为D630大于0.383。特异性和重复性试验表明,建立的间接ELISA方法具有良好的特异性和敏感性,可用于副猪嗜血杆菌的检测。
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| 关 键 词： | 副猪嗜血杆菌 ompP2基因 克隆 表达 间接ELISA |
Cloning and expression of Haemoiphilus parasuis ompP2 gene and development of an indirect enzyme-linked immunosorbent assay for detecting antibody against Haemophilus parasuis |
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| JIA Ai-qing,LI Chun-ling,WANG Gui-ping,HE Yong-long,YUAN Zi-yan.Cloning and expression of Haemoiphilus parasuis ompP2 gene and development of an indirect enzyme-linked immunosorbent assay for detecting antibody against Haemophilus parasuis[J].Chinese Journal of Veterinary Science,2011,31(9). |
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| Authors: | JIA Ai-qing LI Chun-ling WANG Gui-ping HE Yong-long YUAN Zi-yan |
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| Institution: | JIA Ai-qing1,LI Chun-ling2,WANG Gui-ping1,HE Yong-long1,YUAN Zi-yan1 (1.Guangdong Institute of Modern Agricultural Group,Guangzhou 510630,China,2.Institute of Veterinary Medicine Guangdong Academy of Agricultural Science,Guangzhou 510640,China) |
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| Abstract: | The ompP2 gene which encode Haemophilus parasuis outer membrane protein was amplified by PCR.Sequencing results showed that the amplified ompP2 gene was 1 188 bp fragment,the gene was inserted into the expression vector pET-32a.A fusion protein was expressed in BL21 that transfected by pET-32a-ompP2 and induced by IPTG.The molecular weight of the recombinant was about 60 000 by SDS-PAGE,and the immunoreaction activity of the recombinant protein was confirmed by Western-blot.An indirect enzyme-linked immunos... |
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| Keywords: | Haemophilus parasuis ompP2 gene cloning expression indirect ELISA |
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