禽偏肺病毒C型核衣壳N蛋白的哺乳动物细胞表达系统表达及鉴定  

Mammalian expression and identification of avian metapneu movirus subgroup C nucleocapsid protein

作　　者：李子璇[1,2] 韦莉[2] 王菁[2] 朱姗姗[2] 张春燕[2] 柳舒航 全荣[2] 阎旭[2] 陈武[1] 刘爵[2] 

机构地区：[1]北京农学院动物科学技术学院,北京102206 [2]北京市农林科学院畜牧兽医研究所,北京100097

出　　处：《北京农学院学报》2014年第4期49-53,共5页Journal of Beijing University of Agriculture

基　　金：国家现代农业产业技术体系(CARS-42)

摘　　要：利用哺乳动物细胞表达系统对禽偏肺病毒C型核衣壳N蛋白进行表达,并检测所表达融合蛋白的抗原性。首先将N基因克隆到真核表达载体pEGFP-C1中,构建重组真核表达载体,然后将阳性重组质粒转染BHK-21细胞,通过倒置荧光显微镜和共聚焦显微镜检测N蛋白的表达,并用Western blot检测表达蛋白的抗原性。结果显示,禽偏肺病毒核衣壳N蛋白主要表达在细胞浆中,在转染后48h后表达量最高;与抗N蛋白抗体有抗原性,所表达核衣壳N融合蛋白大小为72kDa左右,与预期结果一致。结论,本试验成功应用哺乳动物表达系统表达了禽偏肺病毒C型核衣壳N蛋白,表达蛋白具有抗原性,为与禽偏肺病毒核衣壳N蛋白的相关研究奠定了基础。The aims of this paper were to express subgroup C avian metapneumovirus virus （aMPV） nucleocapsid （N） protein by Eukaryotic expression system and to further analyze the antigenicity of the recombinant protein.The N gene was cloned into eukaryotic expression pEGFP-C1 vector to obtain the recombinant expression vector.The positive recombinant expression vector was transfected into BHK-21 cells.The recombinant N protein expression was analyzed by confocal microscopy and inverted fluorescence microscope and the antigenicity of the recombinant protein by western-blotting.The results demonstrated that the N protein expressed mainly in the cytoplasm and exhibited a specific and single band with a size of about 72 kDa when reacted with a monoclonal antibody raised against subgroup C aMPV N protein.In conclusion,this study expressed aMPV N protein in mammalian expression system with a good antigenicity,which made the foundation work for the relatied study of aMPV N protein.

关 键 词：禽偏肺病毒核衣壳N蛋白 哺乳动物细胞表达系统 显微镜观察 WESTERN BLOTTING 

分 类 号：S851.2[农业科学—预防兽医学]