Expression of cDNA for recombinant human granulocyte colony-stimulating factor inEscherichia coli and characterization of the protein

Objective: To determine the biological activity of rhG-CSF and it’s characterization. Methods: The prokaryotic expression vector pG01 containing human G-CSF cDNA were constructed with DNA recombination technology. Results: We had achieved high level expression of the human G-CSF inE. coli, where it represented at least 23.6% of the total protein as determined from SDS-PAGE gels. The human G-CSF was expressed as inclusion bodies inE.coli. The inclusion bodies were solubilized in a solution containing 7M urea, renatured by dialysis, isolated and purified by DEAE-sepharose CL-6B ion exchange and Superdex 75 gel filtration chromatography. The purified rhG-CSF was confirmed by coincidence of biological activity and protein demonstrated by SDS-PAGE. It was homogeneous with respect to mol. Wt (18400). The purity of the rhG-CSF might be >90 per cent. Conclusion: The purified rhG-CSF in our laboratory had dramatically the biological activity of regulating proliferation and differentiation of the human G-CSF-dependent cell line NSF-1 and the progenitor cells of granulocytes of human bone marrow.