稳定表达猪瘟病毒E2蛋白的BHK细胞系的构建

Construction of BHK Cell Line Stably Expressing E2 Protein of Classical Swine Fever Virus

关键词:  猪瘟病毒  E2蛋白  慢病毒  BHK-21细胞

Key words:  classical swine fever virus  E2 protein  lentivirus  BHK-21 cells

摘要:

猪瘟病毒(CSFV)的E2蛋白是引起猪体产生针对猪瘟保护性抗体的主要抗原，构建可稳定表达CSFV E2蛋白的细胞系，可为E2蛋白功能研究及基因工程猪瘟疫苗的研制提供物质基础。本研究将CSFV E2基因克隆至慢病毒表达载体，构建重组慢病毒表达质粒pCDH-E2。将pCDH-E2重组质粒与慢病毒包装质粒共转染293T细胞，获得重组慢病毒。将该慢病毒感染BHK-21细胞，经嘌呤霉素抗性筛选结合有限稀释法筛选出可表达CSFV E2蛋白的BHK细胞系。Western blot分析结果显示，所构建的重组细胞系传至第10代仍能稳定表达CSFV E2蛋白。该细胞系的建立为研制猪瘟新型重组疫苗及其生产奠定基础。

Abstract:

E2 protein of classical swine fever virus (CSFV) is the main antigen that causes swine to produce protective antibodies against CSFV. Construction of cell lines that can stably express CSFV E2 protein can provide material basis for the research of E2 protein function and genetic engineering vaccine. In this study, CSFV E2 gene was cloned into lentiviral expression vector to obtain the recombinant plasmid of pCDH-E2, then the positive plasmid pCDH-E2 was transfected into 293T cells together with the packaging plasmids to get the recombinant lentivirus. BHK-21 cells were infected with the recombinant lentivirus, and screened by puromycin combined with limited dilution method to establish the cell line expressing CSFV E2 protein. Western blot analysis showed that the recombinant cell line could still express CSFV E2 protein stably in the 10th generation. This study successfully constructed a cell line which can stably express CSFV E2 protein, it laid a foundation for the development and production of new recombinant classical swine fever vaccine.