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| 小反刍兽疫病毒分离株China/Tib/07辅助质粒及微型基因组的构建和鉴定 | |
| 引用本文： | 刘文华,王志亮,包静月,吴晓东,刘华雷,赵永刚,刘秀梵.小反刍兽疫病毒分离株China/Tib/07辅助质粒及微型基因组的构建和鉴定[J].畜牧兽医学报,2011,42(10). |
| 作者姓名： | 刘文华 王志亮 包静月 吴晓东 刘华雷 赵永刚 刘秀梵 |
| 作者单位： | 1. 扬州大学兽医学院,扬州225009;中国动物卫生与流行病学中心国家外来动物疫病诊断中心,青岛266032;青岛农业大学动物科技学院,青岛266109 2. 中国动物卫生与流行病学中心国家外来动物疫病诊断中心,青岛,266032 3. 扬州大学兽医学院,扬州,225009 |
| 基金项目： | 农业部948项目(2006-G57 |
| 摘 要： | 本研究旨在构建小反刍兽疫病毒(PPRV)的辅助质粒和微型基因组并验证其功能,为建立PPRV反向遗传操作平台奠定基础.首先构建表达PPRV分离株China/Tib/07的N、P和L蛋白的辅助质粒pCI-N、pCI-P和pCI-L.通过间接免疫荧光试验验证3个辅助质粒转染细胞后的蛋白表达情况,并从mRNA水平上验证蛋白表达的正确性；扩增PPRV基因组两末端序列和eGFP报告基因,三者通过overlap-PCR连接并克隆至转录载体pOL-TV5中构建微型基因组.将构建的微型基因组分别与辅助病毒和3个辅助质粒进行共转染.经鉴定各辅助质粒完全正确,构建的微型基因组不论与辅助病毒还是与3个辅助质粒共转染,报告基因均表达.本试验成功构建了PPRV分离株的表达N、P和L蛋白的3个辅助质粒以及微型基因组,并用微型基因组验证了3个辅助质粒可以作为PPRV反向遗传操作的辅助质粒,为该病毒感染性克隆的构建奠定了基础. |
| 关 键 词： | 小反刍兽疫病毒 微型基因组 eGFP N蛋白 P蛋白 L蛋白 |
Construction and Identification of Support Plasmids and Minigenome of a Peste des Petits Ruminants Virus Isolate, China/Tib/07 |
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| LIU Wen-hua,WANG Zhi-liang,BAO Jing-yue,WU Xiao-dong,LIU Hua-lei,ZHAO Yong-gang,HU Xiu-fan.Construction and Identification of Support Plasmids and Minigenome of a Peste des Petits Ruminants Virus Isolate, China/Tib/07[J].Acta Veterinaria et Zootechnica Sinica,2011,42(10). | |
| Authors: | LIU Wen-hua WANG Zhi-liang BAO Jing-yue WU Xiao-dong LIU Hua-lei ZHAO Yong-gang HU Xiu-fan |
| Institution: | LIU Wen-hua1,2,3,WANG Zhi-liang2*,BAO Jing-yue2,WU Xiao-dong2,LIU Hua-lei2,ZHAO Yong-gang2,LIU Xiu-fan1 (1.College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China,2.National Diagnostic Center for Exotic Animal Diseases,China Animal Health and Epidemiology Center,Qingdao 266032,3.Animal Science and Veterinary Medicine College,Qingdao Agricultural University,Qingdao 266109,China) |
| Abstract: | The research was conducted to construct and identify the support plasmids and minigenome of Peste des petits ruminants virus(PPRV) in order to lay a foundation for the establishment of a reverse genetic system for rescue of PPRV.Support plasmids of pCI-N,pCI-P and pCI-L according to the PPRV isolate,China/Tib/07,were constructed and functions were identified by IFA and RT-PCR after transfection.At the same time,the two end sequences of the genome of the PPRV isolate,along with the reporter gene of eGFP were... |
| Keywords: | Peste des petits ruminants virus minigenome eGFP N protein P protein L protein |
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