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H9N2亚型禽流感病毒病毒样颗粒的构建
王粲,阮志华,谢芝勋,李孟,罗思思,张民秀,李丹,谢丽基,谢志勤
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(广西壮族自治区兽医研究所/广西兽医生物技术重点实验室/农业农村部中国广西 -东盟跨境动物疫病防控重点实验室)
摘要:
为构建H9N2亚型禽流感病毒病毒样颗粒,本研究以血凝素(Hemagglutinin,HA)、神经氨酸酶(Neurami-nidase,NA)及基质蛋白(Matrix protein,M1)3种蛋白为对象,通过Bac-to-Bac杆状病毒表达系统构建了2种重组杆状病毒,即rBV-HA-M1和rBV-NA,用IFA和Western-blot鉴定重组蛋白的免疫活性,再将rBV-HA-M1和rBV-NA以最佳感染复数比例共感染昆虫Sf9细胞,经浓缩后获得毒样颗粒,同时进行血凝滴度测定和透射电镜观察。透射电镜观察结果显示,表达的HA-M1和NA重组蛋白可以在Sf9细胞中自组装形成病毒样颗粒,浓缩的病毒样颗粒能够凝集1%的鸡红细胞,血凝效价为1:64。试验结果为进一步研究H9N2亚型禽流感病毒病毒样颗粒疫苗奠定了基础。
关键词:  禽流感病毒  H9N2亚型  病毒样颗粒  Bac-to-Bac杆状病毒表达系统
DOI:
投稿时间:2023-11-11修订日期:2024-07-05
基金项目:广西重点研发计划禽流感综合防控技术研究与应用(桂科AB21076004); “广西八桂学者”项目(2019A50);广西重大专项项目(桂科AA23062050);广西科技先锋队项目(桂农科盟202409-01)
Construction of Virus-Like Particles of H9N2 Avian Influenza Virus
(Key Laboratory of China (Guangxi)-ASEAN Cross-border Animal Disease Prevention and Control, Ministry of Agriculture and Rural Affairs, Guangxi Key Laboratory of Veterinary Biotechnology, Guangxi Veterinary Research Institute)
Abstract:
In order to construct virus-like particles of H9N2 avian influenza virus, this study used the proteins of hemagglutinin (HA), neuraminidase (NA) and matrix protein (M1) to construct two recombinant baculoviruses rBV-HA-M1 and rBV-NA, through the Bac-to-Bac baculovirus expression system. The immune activity of the recombinant proteins were identified by IFA and Western-blot, then the Sf9 insect cells were co-infected with rBV-HA-M1 and rBV-NA at the optimal infection complex ratio. The co-infected Sf9 cells were concentrated to obtain the virus-like particles. The virus-like particles were measured for hemagglutination titers and transmission electron microscopy. The results of transmission electron microscopy showed that the expressed HA-M1 and NA recombinant proteins could self-assemble to form virus-like particles in Sf9 cells. The virus-like particles were able to agglutinate 1% of chicken red blood cells, and the titer was 1:64. The results of this study lay the foundation for further research on the virus-like particle vaccine of H9N2 virus.
Key words:  Avian influenza virus  subtype H9N2  virus-like particles  Bac-to-Bac baculovirus expression system

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