稳定转染CD151或CD163的Marc145细胞系的建立及其在PRRS疫苗生产中的应用  

Establishment of Marc145 cell lines stably transfected with CD151 or CD163 and the application in the production of PRRS vaccine

作　　者：阮征 刘开武 吴建英 华娟 夏瑜 王莲芳 胡小明[3] 刘杰[1,3] 黄海军 

机构地区：[1]武汉市农科院畜牧兽医科学研究所,湖北武汉430208 [2]武汉市重大动物疫病防控中心,湖北武汉430023 [3]华中农业大学农业动物遗传育种与繁殖教育部重点实验室,湖北武汉430070

出　　处：《中国兽医杂志》2015年第6期93-96,I0006,共5页Chinese Journal of Veterinary Medicine

基　　金：国家自然科学基金项目(31201938);武汉市科技局项目(2013020705070350-9;-14);武汉市农业科学院创新项目(CX201309);武汉市农业科学院青年英才项目(YC201101)

摘　　要：为了进一步探讨提高PRRSV感染靶细胞滴度的有效方法,本研究采用脂质体介导方法,通过G418筛选稳定转染Marc145细胞,并利用单个细胞克隆技术,获得了稳定超表达CD151蛋白和(或)CD163蛋白的Marc145细胞株,经免疫荧光化学和Western Blot鉴定证实,成功表达出基因重组猪CD151蛋白和CD163蛋白。在GMP生产车间将超表达CD151、CD163蛋白或CD151/CD163蛋白共表达的Marc145细胞感染PRRSV,使病毒感染滴度提高了10倍以上。将上述收获的病毒液制备了灭活苗,疫苗效力超过部颁标准。In order to further explore the effective method to improve the PRRSV titer in infected target cells, the Marc145 cell lines stably over-expressing CD 151 and （or） CD 163 protein were established by liposome mediated method following G418 screen- ing and single cell cloning technology. The successful expression of recombinant porcine CD151 and CD163 proteins was identified by fluorescent immunoeytoehemistry and western Blot. Small scale experiment was conducted in the GMP production workshop and, through the over-expression of PRRSV receptor CD151, CD163 or CD151 and CD163 coexpressing, the virus titer in Marc145 cells infected with PRRSV increased by more than 10 times. Finally, the virus was harvested and the inactivated Vaccine was pro- duced. The vaccine ettleacy was qualified and higher than the standard promulgated by the Ministry of Agriculture.

关 键 词：猪繁殖与呼吸综合征 CD151分子 CD163分子 真核表达 疫苗生产 

分 类 号：S852.651[农业科学—基础兽医学]