Differential expression of human basic fibroblast growth factor in Escherichia coli: potential role of promoter

• Published in: World journal of microbiology & biotechnology 2004-03, Vol.20 (2), p.161-165
• Main Authors: MIRZAHOSEINI, Hasan, MEHRAEIN, Farideh, OMIDINIA, Eskandar, RAZAVI, Mohamad R
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; E coli; Enzymes; Escherichia coli; Fibroblasts; Fundamental and applied biological sciences. Psychology; Genes; Genetic engineering; Genetic technics; Growth factors; Methods. Procedures. Technologies; Modification of gene expression level; Proteins
• ISSN: 0959-3993; 1573-0972
• DOI: 10.1023/B:WIBI.0000021750.22050.55

Four different expression systems were developed for expression of the cDNA encoding human basic fibroblast growth factor (hbFGF), using Escherichia coli as host organism. The hbfgf structural gene was cloned into four expression vectors, pET-3a, pTrc99A, pPR37 and pKK223-3 differing only in their promoters, which were T7, trc, λPR and tac respectively. Expression of the gene was induced by adding 0.5 mM (final concentration) of isopropyl-β-D-thio-galactopyranoside (IPTG) for the vectors carrying T7, trc and tac promoters or by a temperature shift from 30 to 42 °C for the vector carrying λPR. The highest level of expression was observed in pET-1005 (a pET-3a derivative)/BL21 (DE3) system with 18.5 mg/l rhbFGF and the second high level expression was in pR37-1007 (pPR37 derivative) BL21 (DE3) system with 5 mg of rhbFGF/l. Since in the latter system a temperature shift was used for induction, 29% of the hbFGF was recovered as inclusion bodies in the insoluble cell fraction. The level of expression for the two other systems (pTrc-1006 and pKK-1008) was very low.[PUBLICATION ABSTRACT]