Gene cloning, expression and purification of fusion protein epidermal growth factor-linker-trichosanthin.
- Author:
Yong-Mei LI
1
;
Hai-Wen YANG
;
Ren LUO
Author Information
- Publication Type:Journal Article
- MeSH: Blotting, Western; Cloning, Molecular; Electrophoresis, Polyacrylamide Gel; Epidermal Growth Factor; genetics; metabolism; Escherichia coli; genetics; Genetic Vectors; genetics; Humans; Recombinant Fusion Proteins; genetics; isolation & purification; metabolism; Trichosanthin; genetics; metabolism
- From: Journal of Southern Medical University 2007;27(2):205-207
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct a recombinant expression vector of the fusion protein epidermal growth factor (EGF)-Linker-trichosanthin (TCS) and achieve its expression in E. coli to obtain purified EGF-linker-TCS fusion protein.
METHODSThe gene fragments of EGF-linker were amplified by PCR and inserted into the expression plasmid PQE30-TCS, followed by transformation of the recombinant plasmid into E. coli M15 for expression of the fusion protein. Ni-FF column chromatography was utilized for purification of the expressed product.
RESULTSThe recombinant plasmid PQE30-EGF-linker-TCS was stably and highly expressed in E. coli M15. The expressed product existed in the form of soluble protein accounting for about 40% of total cellular protein and reached a purity of above 95% after purification with Ni-FF column chromatography.
CONCLUSIONThe recombinant plasmid PQE30/EGF-linker-TCS has been successfully constructed, which provides a basis for further structural and functional study of EGF and TCS and their potential clinical application for cancer therapy.
