[A preliminary study on the expression and biological function of recombinant human SCF-TPO fusion protein].
OBJECTIVE
To study the expression of recombinant human SCF-TPO fusion protein and its biological function.


METHODS
Four 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.


RESULTS
The 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.


CONCLUSIONS
Expressed SCF-TPO fusion protein after renaturation has biological activity in promoting the proliferation of MO7e cells.
Liu
Nan
N
Department of Immunology, Affiliated Hospital of AMMS. Laboratory of Immunoassary of NCBA, Beijing 100039, China.
Xi
Yong-zhi
YZ
Guo
Si-qi
SQ
Sun
Yu-ying
YY
Yuan
Zhi-hong
ZH
Cui
Jian-wu
JW
Xi
Cai-xia
CX
Liang
Fei
F
Kong
Fan-hua
FH
D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics / Q000502:physiology; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019089:Stem Cell Factor / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D013926:Thrombopoietin / Q000235:genetics* / Q000378:metabolism / Q000502:physiology
D011993:Recombinant Fusion Proteins; D019089:Stem Cell Factor; D013926:Thrombopoietin