摘要
将猪繁殖与呼吸综合征病毒CH_la株囊膜糖蛋白 (GP5 )基因定向亚克隆到杆状病毒转移载体pBlueBacHisB中 ,与杆状病毒线性DNA(Bac_N_BlueTMDNA)共转染昆虫细胞sf9,经过三轮蚀斑纯化后 ,获得重组杆状病毒rBac_GP5。用该重组病毒感染sf9细胞后 ,应用间接免疫荧光试验、SDS_PAGE和Westernblot检测表明 ,GP5基因在杆状病毒系统中获得了高效表达 ,表达产物因糖基化程度差异 ,表观分子量有 2 2、2 5和 30kDa三种总计约占细胞蛋白总量的 2 6 .4%。
Porcine reproductive and respiratory syndrome virus(PRRSV)strain CH_la,a Chinese reference strain,was isloated in Beijing suburb in the middle 1990 Its GP5 gene encoded by open reading frame(ORF)5 was cloned and inserted into the baculovirus transfer vector pBlueBacHis B. Sf9 cells were co_transfected by the recombinant transfer plasmid and linear baculovirus DNA.A recombinant baculovirus was obtained after blue_plaque screening and three cycles of plaque_purification.The GP5 gene was confirmed to be expressed in sf9 cells by immunofluorescent staining and Western blot.The expressed recombinant fusion protein with Histag have apparent molecular weight of 22,25,and 30,accounting for 26.4% approximately in whole cellular proteins.
出处
《中国预防兽医学报》
CAS
CSCD
2000年第4期278-281,共4页
Chinese Journal of Preventive Veterinary Medicine
基金
国家重点基础研究发展规划项目 !(编号 :G19990 1190 2 )资助
关键词
杆状病毒表达系统
GP5基因
PRRS
重组疫苗
porcine reproductive and respiratory syndrome virus
baculovirus expression system
GP5 gene
