农业生物技术学报2025，Vol.33Issue(5)：1141-1148,8.DOI:10.3969/j.issn.1674-7968.2025.05.017
马麝出血症病毒VP60蛋白在昆虫细胞-杆状病毒系统中的表达与鉴定
Expression and Identification of the VP60 Protein of Moschus chrysogaster hemorrhagic disease virus in the Insect Cell-Baculovirus System
摘要
Abstract
Moschus chrysogaster viral hemorrhagic disease(McVHD)is an acute and highly lethal infectious disease caused by Moschus chrysogaster hemorrhagic disease virus(McHDV)in alpine musk deer.Currently,there are no preventive or therapeutic measures available for this disease.In order to develop a safe and effective vaccine against McVHD,in this study,the full-length gene of McHDV VP60 was amplified by PCR.The VP60 gene was then inserted into the vector pFastBac1,followed by transformation into DH10Bac competent cells containing the Bacmid plasmid.A recombinant bacmid,rBacmid-McHDV-VP60,carrying the VP60 gene,was constructed.Subsequently,Sf9 insect cells were transfected with the recombinant bacmid to obtain the recombinant baculovirus rBac-McHDV-VP60.The expression of the target protein in the recombinant baculovirus rBac-McHDV-VP60 was identified by Western blot and SDS-PAGE analyses.The VP60 protein was identified by transmission electron microscopy and erythrocyte agglutination assay.The results indicated that the bacmid rBacmid-McHDV-VP60 was successfully constructed and the recombinant baculovirus rBac-McHDV-VP60 was obtained.SDS-PAGE and Western blot confirmed that Sf9 cells infected with the recombinant baculovirus produced a soluble McHDV-VP60 protein,approximately 63 kD in size.Transmission electron microscopy showed that the McHDV-VP60 protein self-assembled into virus-like particles(VLPs),which were approximately 35 nm in size,spherical in shape,and smooth on the surface.Erythrocyte agglutination assay showed that the recombinant McHDV-VP60 protein solution had a hemagglutination titer of 1∶215 against 1％human(Homo sapiens)O erythrocytes.In this study,a recombinant baculovirus expressing McHDV-VP60 protein was successfully constructed using an insect cell-baculovirus expression system,and the McHDV-VP60 protein could self-assemble into VLPs.This study provides technical support for the development of a vaccine for McVHD virus-like particles.关键词
马麝出血症病毒(McHDV)/VP60蛋白/重组杆状病毒/病毒样颗粒(VLPs)Key words
Moschus chrysogaster hemorrhagic disease virus(McHDV)/VP60 protein/Recombinant baculovirus/Virus-like particles(VLPs)分类
畜牧业引用本文复制引用
刘玉东,张志雄,邢小勇,包世俊,邵钰,杨永宁,王青,黄冬,何肖肖,王丽,马海云,赵云海..马麝出血症病毒VP60蛋白在昆虫细胞-杆状病毒系统中的表达与鉴定[J].农业生物技术学报,2025,33(5):1141-1148,8.基金项目
甘肃省优秀博士生资助项目(23JRRA1445) （23JRRA1445）
甘肃省重点研发计划—农业类(20YF3NA015) （20YF3NA015）
甘肃农业大学公招博士科研启动基金(GAU-KYQD-2023-25) （GAU-KYQD-2023-25）
国家自然科学基金(32072863) （32072863）
