A preliminary study on the expression and biological function of recombinant human SCF-TPO fusion protein.
- Author:
Nan LIU
1
;
Yong-zhi XI
;
Si-qi GUO
;
Yu-ying SUN
;
Zhi-hong YUAN
;
Jian-wu CUI
;
Cai-xia XI
;
Fei LIANG
;
Fan-hua KONG
Author Information
- Publication Type:Journal Article
- MeSH: Blotting, Western; Cell Line, Tumor; Cell Proliferation; Cell Survival; genetics; physiology; Cloning, Molecular; Electrophoresis, Polyacrylamide Gel; Escherichia coli; genetics; Genetic Vectors; Humans; Recombinant Fusion Proteins; genetics; metabolism; physiology; Reverse Transcriptase Polymerase Chain Reaction; Stem Cell Factor; genetics; metabolism; physiology; Thrombopoietin; genetics; metabolism; physiology
- From: Chinese Journal of Hematology 2005;26(1):19-22
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo study the expression of recombinant human SCF-TPO fusion protein and its biological function.
METHODSFour primers were designed according to known sequences of TPO and SCF. The functional amino acid domains of TPO and SCF were amplified by RT-PCR from fetus hepatocytes, respectively. The expression plasmid pET32a/SCF-TPO was constructed by VOE gene fusion technique and expressed in E. coli BL21(DE3) plysS as inclusion body after isopropyl-beta-D-1-thiogalactopyranoside (IPTG) induction. The fusion protein was tested by SDS-PAGE and Western blot. The biological functions of SCF-TPO fusion protein in MO7e cells was investigated by MTT method after purification with metal chelating chromatography.
RESULTSThe high expression SCF-TPO fusion protein was obtained, reaching up to 30% of the total cellular protein. Western blot verified the correct fusion expression and MTT results showed the growth promoting effect of the SCF-TPO fusion protein on MO7e cells, with a higher promoting activity at 100 ng/ml.
CONCLUSIONSExpressed SCF-TPO fusion protein after renaturation has biological activity in promoting the proliferation of MO7e cells.
