摘要
为鉴定猪繁殖与呼吸综合征病毒SCM株(PRRSV-SCM)囊膜糖蛋白GP5的免疫原性,利用RT-PCR方法从PRRSVSCM株ORF5中扩增大小为231bp的目的基因并构建表达载体PET32-GP5,将其转入大肠埃希菌Rosetta,运用IPTG通过条件优化诱导其表达.检测结果表明:最佳诱导条件为IPTG浓度1mmol/L在37℃下作用5h,表达大小约28kU的融合蛋白.通过Western-blot和间接酶联免疫吸附试验检测表达蛋白,结果显示该原核表达的融合蛋白可与PRRSV阳性血清发生反应,证明该蛋白具有免疫反应性.这为将来研制检测试剂盒以及后续研究PRRSV亚单位疫苗提供了重要依据.
In order to identify the immunogenicity of envelope glycoprotein GP5 of porcine reproductive and respiratory syndrome virus SCM strain（PRRSV-SCM）,the 231 bp target gene was amplified from the ORF5 of the strain PRRSV SCM by RT-PCR method and cloned into expression vector pET32.Then the authors transformed the recombinant plasmid PET32-GP5 into E.coli Rosetta and inducted it to express under the IPTG.The detecting results showed that fusion protein,weight 28 kU,was expressed under the best conditions when IPTG concentration was 1 mmol/L at 37 ℃ for 5 h.Detection of expressed protein by western-blot and ELISA showed that the prokaryotic expression of the fusion protein could react with the positive serum samples of PRRSV.The result indicated that the protein had immune reactivity,which provided an important basis to developed PRRSV test kit in the future as well as the later study of PRRSV subunit vaccine.
出处
《四川师范大学学报（自然科学版）》
CAS
CSCD
北大核心
2010年第5期679-683,共5页
Journal of Sichuan Normal University（Natural Science）
基金
国家"十一五"科技支撑计划基金(2006BAD06A18)
"长江学者和创新团队发展计划"创新团队项目基金(IRT0848)资助项目
