猪繁殖与呼吸综合征病毒的纯化与形态观察  

Purification and morphological observation of porcine reproductive and respiratory syndrome virus

作　　者：魏德陇 张婧[1] 李坤[1] 孙普[1] 白兴文[1] 包慧芳[1] 李冬[1] 刘在新[1] 卢曾军[1] 曹轶梅[1] WEI De-long;ZHANG Jing;LI Kun;SUN Pu;BAI Xing-wen;BAO Hui-fang;LI Dong;LIU Zai-xin;LU Zeng-jun;CAO Yi-mei(State Key Laboratory of Veterinary Etiological Biology/National Foot-and-mouth Disease Reference Laboratory/Key Laboratory of Animal and Poultry Virology of the Ministry of Agriculture/Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Lanzhou 730046,China)

机构地区：[1]中国农业科学院,兰州兽医研究所,家畜疫病病原生物学国家重点实验室,国家口蹄疫参考实验室农业部畜禽病毒学重点开放实验室,甘肃兰州730046

出　　处：《中国兽医科学》2021年第12期1519-1524,共6页Chinese Veterinary Science

基　　金：国家重点研发计划项目(2017YFD0501104);甘肃省青年科技基金项目(20JR10RA022)。

摘　　要：为制备猪繁殖与呼吸综合征病毒(PRRSV)全病毒抗原,用于免疫学研究,本研究在Marc-145细胞上繁殖了PRRSV基因Ⅱ型VR2332株,经聚乙二醇浓缩后,用蔗糖密度梯度离心法纯化PRRSV病毒粒子。用电子显微镜观察病毒粒子形态结构,并用SDS-PAGE、免疫印迹等方法分析了纯化效果。结果显示,电子显微镜观察到PRRSV形状呈球形或椭圆形,囊膜表面有微绒毛样突起,病毒粒子大小不一,大多数直径在50~80 nm之间,有极少数直径超过120 nm。SDS-PAGE可见病毒的多个结构蛋白条带,利用感染康复猪血清进行免疫印迹检测显示,M蛋白的印迹条带最深,显示纯化病毒具有良好的反应活性,为后续免疫学研究奠定了基础。In order to prepare porcine reproductive and respiratory syndrome virus(PRRSV)whole virus antigen for immunological study,VR2332 strain of PRRSV geneⅡwas propagated on Marc-145 cells.After concentrated by polyethylene glycol,PRRS virions were purified by sucrose density gradient centrifugation method.The morphology and structure of virions were observed by electron microscope,and the purification effect was analyzed by SDS-PAGE and Western blot.The results showed that PRRS virions were spherical or elliptical in shape,and there were microvilli on the surface of the envelope.The sizes of the virions were different,most of which were ranging between 50 nm and 80 nm in diameter,and a few were more than 120 nm in diameter.Multiple structural protein bands of the virus can be seen in SDS-PAGE.Western-blot analysis of the viral proteins included in purified whole viruses showed that the M protein had the strongest blotting bands recognized by the serum from PRRSV infected rehabilitated pigs,which indicated that the purified virus had good reactivity and laid a foundation for subsequent immunological studies.

关 键 词：猪繁殖与呼吸综合征病毒 纯化 形态学 蔗糖密度梯度 

分 类 号：S852.651[农业科学—基础兽医学]