首页|猪圆环病毒2d型病毒样颗粒疫苗的制备及发酵工艺研究

猪圆环病毒2d型病毒样颗粒疫苗的制备及发酵工艺研究

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  • 国家科技期刊平台
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为制备猪圆环病毒2d型(PCV2d)病毒样颗粒(VLPs)疫苗并对其免疫效果进行评价.本研究采用原核表达系统表达PCV2d Cap蛋白,并对其发酵工艺条件进行了优化,去除His-SUMO标签使其体外自组装形成VLPs,通过SDS-PAGE和Western-blot进行验证,采用透射电镜和纳米粒度仪进行表征,将验证正确的VLPs免疫小鼠,测定其抗体水平和细胞因子水平.结果显示,在0.5 mmol/L IPTG、N源补料速率为12％时,蛋白表达量最高,利用透射电镜、纳米粒度仪进行表征鉴定,发现PCV2d可自组装形成VLPs,粒子直径约为20～25 nm.PCV2dVLPs与ISA 201佐剂混合免疫小鼠,特异性抗体滴度达到1∶512以上,中和抗体滴度达到1∶32以上,IL-4和IFN-γ细胞因子水平均可达到80 μg/mL,且明显高于PBS组.综上所述,本研究利用原核表达系统构建表达并成功获得PCV2dVLPs,且可以诱导机体产生良好的细胞免疫和体液免疫,这为猪圆环病毒2d型疫苗及猪圆环病毒多价疫苗的研制奠定了基础.
Study on preparation and fermentation technology of porcine circovirus 2d virus-like particles vaccine
To develop a porcine circovirus type 2d virus-like particles vaccine and evaluate its immunization effect,the PCV2d Cap protein were expressed by prokaryotic expression system and confirmed through SDS-PAGE and Western-blot analysis.When the prime conditions of fermentation process were screened,the His-SUMO tag of expressed PCV2d Cap was removed to make its self-assembly into VLPs in vitro.The morphological characterization of the VLPs was checked by transmission electron microscopy and nanoparticle-size analyzer.The antibody and cytokine levels in sera were detected at 28 d after mice immunized with the VLPs.The results showed that the highest level of protein expression was achieved at 0.5 mmol/L IPTG and a 12％N source feed rate,and the VLPs were found to have a particle diameter of approximately 20-25 nm.The titers of specific antibody and neutralizing antibody were more than 1∶512 and 1∶32 respectively,and the levels of IL-4 and IFN-γ cytokines were 80 μg/mL,which were significantly higher than those in the PBS group.In conclusion,this study successfully con-structed and obtained PCV2d VLPs using the prokaryotic expression system,which induced good cellular and humoral immunity and laid the groundwork for research on the porcine circovirus 2d subtype and porcine circovirus multivalent vaccine.

porcine circovirus 2dvirus-like particlesE.coli expression systemfermentation technology

王运航、吕熙、段梦迪、魏甜、李毅、张雅雅、陈相杨、郭慧琛、孙世琪、白满元、敬晓棋

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榆林学院生命科学学院,陕西榆林 719000

中国农业科学院 兰州兽医研究所 兰州大学动物医学与生物安全学院 动物疫病防控全国重点实验室,甘肃兰州 730046

猪圆环病毒2d型 病毒样颗粒 大肠杆菌表达系统 发酵工艺

2023兰州市人才创业创新项目甘肃省在站博士后专项甘肃省科技重大专项国家自然基金青年基金项目陕西省科技厅重点研发项目陕西省教育厅2023年度重点科学研究计划项目-协同创新中心项目榆林学院研究生创新基金项目国家重点研发计划项目国家重点研发计划项目

2023-RC-323JRRA55423ZD-NA007323011272023-YBNY-14323JY0902022YLYCX122022YFD18006032021YFD1800300

2024

10.16656/j.issn.1673-4696.2024.0152

中国兽医科学
中国农业科学院兰州兽医研究所

中国兽医科学

CSTPCD北大核心
影响因子：0.524
ISSN：1673-4696
年,卷(期)：2024.54(10)
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