摘要
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 in E 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 in E 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
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 in E 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 in E 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
