Document Type: Original Article(s)
Department of Genetic Engineering, Research Center for Science and Biotechnology, Tehran, Iran
Department of Biotechnology, Pasteur Institute of Iran, Tehran, Iran
Department of Hematology, Tarbiat Modarres University, School of Medicine,Tehran, Iran
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.