Adenovirns construction with miceCD40- IgG2aFc-IRS-GFP fusion gene and in vitro testing
10.3760/cma.j.issn.1007-8118.2010.03.017
- VernacularTitle:构建含有小鼠CD40-IgG2aFc-IRS-GFP融合基因腺病毒并体外检测
- Author:
Chuanguang LI
;
Yifeng FANG
;
Yingxia TAN
;
Zhijian SHEN
;
Qiyu ZHANG
- Publication Type:Journal Article
- Keywords:
CD40;
PDC316 plasmid;
Fusion gene;
Plasmid construction;
Reconstructed adenovirus
- From:
Chinese Journal of Hepatobiliary Surgery
2010;16(3):208-211
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the adenovirus vector containing mice CD40- IgG2aFc-IRS-GFP fusion gene and detect the expression of the fusion protein in transfeete 293cells.Methods The fragments of CD40, IgG2aFc were obtained and inserted into plasmid PDC316-IgG2aFc-GFP.After being verified by sequencing, Ad5-PDC316-CD40-IgG2aFc-GFP was contransfored with the bckbone vector into 293 cells.The virus titer was detected after replicating and purifing and the expression of the fusion protein was analyzed.Results A virus and plasmid were constructed successfully.The vitro infection showed that the virus can infect cells of which the fusion protein was confirmed by fluo-rescence.The expression of the fusion protein increased with the increased time and virus concentra-tion.The protein expression stopped increasing after the virus concentration came to a certain level.Conclusion CD40, IgG2aFc fragments are correctly ligated with plasmid PDC316-IgG2aFc-GFP, and the fusion protein can be expressed in 293cells.This might lay a foundation for further studies of the expresstion of virus, immune tolerance and mechanism of liver transplantation model in rats.
