Construction of the eukaryotic expression vector containing rat transforming growth factor beta type II receptor and interferon gamma fusion genes.
- Author:
Zi-yang YU
1
;
Li-huang ZHANG
;
Hang-ping YAO
;
Xiao-hua YANG
Author Information
- Publication Type:Journal Article
- MeSH: Amino Acid Sequence; Animals; Base Sequence; CHO Cells; Cloning, Molecular; Cricetinae; Eukaryotic Cells; metabolism; Genetic Vectors; Hepatocytes; metabolism; Interferon-gamma; biosynthesis; genetics; Molecular Sequence Data; Plasmids; genetics; Protein-Serine-Threonine Kinases; Rats; Receptors, Transforming Growth Factor beta; biosynthesis; genetics; Recombinant Fusion Proteins; biosynthesis; genetics; Recombinant Proteins
- From: Journal of Zhejiang University. Medical sciences 2005;34(1):43-54
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo express a bioactive fusion protein comprising rat soluble transforming growth factor beta type II receptor and interferon gamma(rsTGFbetaR II-IFN gamma) in mammalian cells.
METHODSmRNA was extracted from rat liver and the sTGFbetaR II-IFN gamma genes amplified by RT-PCR, then the two gene segments were cloned into the same pSecTag2A expression vector, and pSecTag2A/rsTGFbetaR II-IFN gamma recombinant plasmid was obtained, which was later transfected into CHO cells using liposomes. The expression of pSecTag2A/rsTGFbetaR II-IFN gamma in the supernatant was detected by ELISA and Western blotting. The bioactivities of the fusion protein were tested by sTGF betaR II-IFN gamma and IFN gamma bioassays.
RESULTSpSecTag2A/rsTGFbetaR II-IFN gamma transfectants expressed rsTGF betaR II-IFN gamma fusion protein. The purified fusion protein exhibited anti-viral activity and antagonized the proliferation-inhibitive effect of TGFbeta1 on CCL-64 cells. It inhibits the HSC activation in vitro.
CONCLUSIONThe pSecTag2A/rsTGFbetaR II-IFN gamma recombinant plasmid constructed in this study can express bioactive rsTGFbetaR II-IFN gamma fusion protein.
