Cloning, expression of human keratinocyte growth factor and its purification and identification.
- Author:
Bin-Wen WU
1
;
Zhao-Jun DUAN
;
Wu-Ping LI
;
Yong CHEN
;
Hong-Liang LÜ
;
Zuo-An YI
;
Cheng-Hai ZHANG
;
Ju-Sheng LIN
;
Jia-Long WANG
;
Yun-De HOU
Author Information
1. National Laboratory of Molecular Virology and Genetic Engineering, Institute of Virus Prevention and Control, Chinese Center of Disease Control, Beijing 100052, China.
- Publication Type:Journal Article
- MeSH:
Cloning, Molecular;
Escherichia coli;
genetics;
metabolism;
Fetus;
Fibroblast Growth Factor 10;
biosynthesis;
genetics;
isolation & purification;
Genetic Vectors;
genetics;
Humans;
Lung;
chemistry;
Recombinant Proteins;
biosynthesis;
genetics;
isolation & purification
- From:
Chinese Journal of Biotechnology
2004;20(3):461-464
- CountryChina
- Language:Chinese
-
Abstract:
To clone KGF-2 gene, get hKGF-2 protein and detemine its activity. The cNDA of human KGF-2 was isolated from fetal lung by RT-PCR and cloned into pBV220 plasmid. The recombinant pBV220-hKGF-2 plasmid was transformed into E. coli (BL21), induced at 42 degrees C for the expression of hKGF-2. Recombinant human KGF-2 was purified from the ultrasonic-treated BL21 by heparin-Sepharose CL-6B treated column chromatography and cation exchange column chromatography. MTT method was used for the determination of its biological activity. SDS-PAGE showed that rhKGF-2 was expressed in E. coli BL21 as soluble protein of approximately 20kD. The rhKGF-2 protein can stimulate the proliferation of NIH3T3 cells significantly from 1 ng/mL to 10 ng/mL. HKGF-2 cDNA wasclned and highly expressed in E. coli BL21 and the purified rhKGF-2 showed the mitogenic activity on NIH3T3 cells.
