尼帕病毒和亨得拉病毒核蛋白基因在杆状病毒表达系统的表达及反应原性鉴定  

The Expression of Nucleocapsid Protein of Nipah and Hendra Viruses in Baculovirus and Characterization of Their Reactivity

作　　者：肖昌[1] 徐兴然[1] 查云峰[1] 张玉静[2] 宣华[3] 涂长春[1] 

机构地区：[1]军事医学科学院军事兽医研究所病毒室,吉林长春130062 [2]吉林大学农学部畜牧兽医学院,吉林长春130062 [3]广东省农业科学院兽医研究所,广东广州510640

出　　处：《病毒学报》2006年第6期466-470,共5页Chinese Journal of Virology

基　　金：科技部重大动物疫病防治平台项目(2004BA519A48);广东省科技计划项目(2004A20403001)

摘　　要：利用改造的昆虫杆状病毒表达系统转移载体，成功地对尼帕病毒（NiV）和亨得拉病毒（HeV）的核蛋白基因进行了分泌表达。重组蛋白表达的时效性分析结果显示，Sf9细胞感染重组病毒72～96h后，蛋白的表达量达到高峰，大规模表达可分别获得2．3～2．4mg／L纯化的目的蛋白。应用Western blot、间接ELISA和间接免疫荧光（IFA）对两种重组N蛋白与兔抗NiV和HeV阳性血清的反应性进行鉴定，结果表明两种重组N蛋白和阳性血清有很好的反应性，并且在血清学上有交叉反应。该研究为今后开展流行病学调查和诊断试剂的研制打下了基础。Nipah virus（NiV） and Hendra virus（HeV）, as two novel members of Henipah genus in paramyxoviridae, are the causative agents of fetal Nipah and Hendra zoonoses, which first emerged in 1998 in Malaysia and in 1994 in Australia. These diseases feature the severe pathogenic damages on respiratory system and central nervous system, usually associated with high morbidity and mortality in host animals and the human. In order to prevent their potential outbreak in China, in this study the NiV N and HeV N genes of the viruses were amplified by PCR from the plasmids imported from Australia, and cloned into the pFastBac-Mels-6His vector separately, resulting in the expression in baculovirus system of N protein fused with His-tag at its C terminus. The expression of N proteins in Sf9 cells showed that the highest level of the protein expression could be achieved at 72--96h postinfection, and about 2.3 - 2.4 mg/L of target proteins could be obtained respectively in large scale expression test. The indirect-ELISA and Western blot results showed that the N proteins expressed could react with rabbit anti-NiV and rabbit anti-HeV sera. The indirect immunofluorescence assay（ IFA） using rabbit anti- NiV and rabbit anti-HeV sera could identify the cells expressing N proteins on Sf9 cell monolayers. Taken together, the results showed that the efficient expression of N protein of both NiV and HeV has been achieved and the resulted proteins can be used in future as diagnostic or preventive reagent.

关 键 词：副粘病毒科 尼帕病毒 亨得拉病毒 核蛋白 杆状病毒 表达 反应原性 

分 类 号：Q786[生物学—分子生物学]