overexpression of recombinant human granulocyte colony-stimulating factor in e. coli

bakground: granulocyte colony-stimulating factor (g-csf) is a cytokine that stimulates hematopoiesis and induces proliferation and differentiation of granulocyte progenitor cells as well as production of bone marrow neutrophilic granulocyte colonies.  nowadays, human recombinant g-csf(hr g-csf) is used for the treatment of chemotherapy- and radiotherapy-induced neutropenia, and also in patients with bone marrow transplantation.   methods: a cdna of human g-csf (hg-csf) was synthesized by pcr from recombinant cloning vector, with two altered nucleotides for increasing mrna stability and overexpression, then inserted into a pet expression vector under the control of t7 promoter and cloned in e. coli strain bl21 (de3).     results: after culture and induction of recombinant e. coli with iptg, we achieved a high level expression of the hg-csf, where it represented approximately 35% of the total protein as determined by sds-page and confirmed by western blotting with polyclonal and monoclonal hg-csf antibodies.   conclusion: rhg-csf was produced in a significantly high quantity with a yield of 35% of total protein as determined by sds-page.  since it is easily obtained by simple purification steps, it may be cost-effective, even on an industrial scale.