Optimization of culture conditions for high- level expression of fibroblast growth factor -2 (FGF- 2) in escherichia coli

Abstract

Fibroblast growth factor-2 (FGF-2), a member of FGF family, is a growth factor which is a critical component in human embryonic stems cell culture medium to maintain the undifferentiated state of stem cells. These undifferentiated stem cells then can develop into any cell types of human body. This unique property will raise a lot of applications of human stem cells in health and medical researches. In Vietnam, however, the cost of FGF-2 is still high, this cause the high cost of human embryonic stem cells for other applications. For that reason, the aim of this thesis is to optimize the best culture conditions for producing FGF-2 with high quality and low cost. In this study, the plasmid pET-FGF was isolated from E. coli DH5α/pET-FGF, which was provided by University of Science, VNU-HCM, and then transformed into the host strain E. coli BL21(DE3). The expression of pET-FGF was induced by IPTG and confirmed by SDS-PAGE and Western blot using specific antibody. After successfully construction of E. coli BL21 (DE3)/pET-FGF, the optimization of induction conditions were conducted via induction temperature, shaking speed, inducer concentration and induction time to select the best conditions. Among the induction conditions that were optimized, the highest level of FGF- 2 expression is achieved at 30°C, with a five hours induction time at 300rpm shaking, using 0.6mM IPTG final concentration as an inducer. This scheme will raise the production of FGF-2 in E. coli strain.