| 日本脑炎病毒NS3基因截短表达及其在神经细胞定位研究 |
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| 引用本文： | 邓绪芳,史子学,魏建超,邵东华,王少辉,李蓓蓓,马志永.日本脑炎病毒NS3基因截短表达及其在神经细胞定位研究[J].中国动物传染病学报,2012(4):7-12. |
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| 作者姓名： | 邓绪芳 史子学 魏建超 邵东华 王少辉 李蓓蓓 马志永 |
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| 作者单位： | 中国农业科学院上海兽医研究所 |
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| 基金项目： | 国家国际科技合作项目(2010DFB33920);中央级公益性科研院所基本科研业务费专项资金(2011JB06) |
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| 摘 要： | 根据GenBank发表的日本脑炎病毒(Japanese encephalitis virus,JEV)基因序列,合成用于克隆编码病毒NS3蛋白C端的基因片段(955~1857 bp)的引物。从感染JEV的细胞样品中,采用RT-PCR扩增出大小约900 bp的目的片段,插入到pET-28a(+)表达载体,诱导表达出NS3C蛋白。将该重组蛋白免疫小鼠制备NS3C抗血清,通过间接免疫荧光和Western blot验证,该抗血清能特异识别JEV感染Vero细胞中的NS3蛋白。使用该抗血清研究了JEV在神经细胞内的亚定位,发现NS3主要定位于JEV感染的神经细胞的胞浆中,并在细胞核周围分布明显。
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| 关 键 词： | 日本脑炎病毒 NS3 细胞定位 |
EXPRESSION OF TRUNCATED NS3 OF JAPANESE ENCEPHALITIS VIRUS AND ANALYSIS OF NS3 SUBCELLULAR LOCALIZATION IN NEUROCYTES |
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| DENG Xu-fang,SHI Zi-xue,WEI Jian-chao,SHAO Dong-hua,WANG Shao-hui, LI Bei-bei,MA Zhi-yong.EXPRESSION OF TRUNCATED NS3 OF JAPANESE ENCEPHALITIS VIRUS AND ANALYSIS OF NS3 SUBCELLULAR LOCALIZATION IN NEUROCYTES[J].Chinese Journal of Animal Infectious Diseases,2012(4):7-12. |
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| Authors: | DENG Xu-fang SHI Zi-xue WEI Jian-chao SHAO Dong-hua WANG Shao-hui LI Bei-bei MA Zhi-yong |
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| Institution: | (Shanghai Veterinary Research Institute,CAAS,Shanghai 200241,China) |
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| Abstract: | Specific primers based on the gene sequence of Japanese encephalitis virus(JEV) deposited in GenBank were synthesized and used to amplify a gene fragment(955-1857 bp) encoding the C terminal of NS3 protein by RT-PCR.The amplified products(approximately 900 bp) were inserted into plasmid pET-28a(+) to express truncated NS3 protein(NS3C).The expressed NS3C was injected into mice to produce antiserum against NS3.Immunofluorescence and Western blot analysis of the specificity of the NS3C antiserum indicated that the NS3C antiserum specifically reacted with NS3 expressed in JEV-infected Vero cells.The subcellular localization of NS3 in JEV-infected neurocytes was detected using the NS3C antiserum.The viral NS3 was found to localize predominantly in the cytoplasm,especially in the perinuclear area,of JEV-infected neurocytes. |
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| Keywords: | Japanese encephalitis virus NS3 cellular localization |
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