Cloning, expression, and purification of a synthetic human growth hormone in Escherichia coli using response surface methodology.

Abstract

The aim of this study was to achieve high-level production of the human growth hormone (hGH) in the prokaryotic expression system. In this regard, we performed cloning, expression, and purification of a synthetic hGH gene in BL21 (DE3) strain of E. coli. The hGH production was determined by SDS-PAGE and western blotting techniques, and then the protein concentration was determined by the Bradford assay. To gain insight into the effect of different nutrients on the growth of E. coli and hGH production, in a preliminary assessment nine different types of the basal medium were analyzed. The highest growth of E. coli and hGH production were observed in TB and SOB media. Accordingly, design of experiments was employed for screening the most significant nutrients, and central composite face design was applied for the optimization. The optimum medium consisted of yeast extract (10 g/L), tryptone (10 g/L), and K2HPO4 (2 g/L). The optimum hGH concentration was 391 mg/L, which was 3-fold higher than the hGH concentration in the LB basal medium (119 mg/L). This production rate is the highest hGH concentration reported in the IPTG-inducible expression systems.

DOI: 10.1007/s12033-014-9818-1

Cite this paper

@article{Zamani2015CloningEA, title={Cloning, expression, and purification of a synthetic human growth hormone in Escherichia coli using response surface methodology.}, author={Mozhdeh Zamani and Aydin Berenjian and Shiva Hemmati and Navid Nezafat and Mohammad Bagher Ghoshoon and Fatemeh Dabbagh and Milad Mohkam and Younes Ghasemi}, journal={Molecular biotechnology}, year={2015}, volume={57 3}, pages={241-50} }