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Information Journal Paper

Title

Production Design of Efficient Recombinant Human Granulocyte-Macrophage Colony-Stimulating Factor Under a Gene-Specific Promoter in Prokaryotic System

Pages

  314-321

Abstract

 Objectives: Recombinant products play an important role in improving health conditions. In addition, human granulocyte-macrophage colony-stimulating factor (hGM-CSF) is considered a cytokine which stimulates many differentiated myeloid cells in order to produce granulocytes, macrophages, and monocytes. Considering the clinical application of the human GM-CSF, the current study aimed to produce the Recombinant human GM-CSF (rhGM-CSF) in the prokaryotic system and then evaluated its biological activity. Materials and Methods: In this experimental study, the hGM-CSF was synthesized under a specific Promoter. Then, it was cloned in HindIII restriction enzyme sites of the pcDNA3. 1 (+). The hGM-CSF gene cloning was assessed by polymerase chain reaction, restriction enzyme digestion, and sequencing. Subsequently, recombinant plasmids were transformed in Escherichia coli and the expression of recombinant hGM-CSF was analyzed by electrophoresis and immunoblotting. Then, the rhGM-CSF was purified using S-tag affinity chromatography and the concentration of the purified rhGM-CSF was determined by ELISA. Finally, the biological function of the rhGM-CSF on TF-1 cells was performed by MTT proliferation assay. Results: The cloned fragment on gel agarose was detected. Further, the restriction enzyme digestion and recombinant plasmid sequencing results confirmed pcDNA3. 1 (+)/hGM-CSF cloning. Furthermore, the results of the expression analysis of rhGM-CSF by SDS-PAGE and western blot showed a specific protein band. The concentration of the purified protein was 0. 54 μ g/mL. Moreover, the proliferation index demonstrated that the treated cells were proliferated (P < 0. 05). The mean values of the proliferation index were 7. 8. Conclusions: In general, the production of recombinant hGM-CSF protein in the prokaryotic system was simple, rapid, and inexpensive. Therefore, the functional rhGM-CSF can be expressed under gene-specific Promoter without any need for the chemical inducer.

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    APA: Copy

    Vahidi, Mahmoud, BANDEHPOUR, MOJGAN, KAZEMI, BAHRAM, & BOZORGMEHR, MAHMOOD. (2020). Production Design of Efficient Recombinant Human Granulocyte-Macrophage Colony-Stimulating Factor Under a Gene-Specific Promoter in Prokaryotic System. CRESCENT JOURNAL OF MEDICAL AND BIOLOGICAL SCIENCES, 7(3), 314-321. SID. https://sid.ir/paper/778550/en

    Vancouver: Copy

    Vahidi Mahmoud, BANDEHPOUR MOJGAN, KAZEMI BAHRAM, BOZORGMEHR MAHMOOD. Production Design of Efficient Recombinant Human Granulocyte-Macrophage Colony-Stimulating Factor Under a Gene-Specific Promoter in Prokaryotic System. CRESCENT JOURNAL OF MEDICAL AND BIOLOGICAL SCIENCES[Internet]. 2020;7(3):314-321. Available from: https://sid.ir/paper/778550/en

    IEEE: Copy

    Mahmoud Vahidi, MOJGAN BANDEHPOUR, BAHRAM KAZEMI, and MAHMOOD BOZORGMEHR, “Production Design of Efficient Recombinant Human Granulocyte-Macrophage Colony-Stimulating Factor Under a Gene-Specific Promoter in Prokaryotic System,” CRESCENT JOURNAL OF MEDICAL AND BIOLOGICAL SCIENCES, vol. 7, no. 3, pp. 314–321, 2020, [Online]. Available: https://sid.ir/paper/778550/en

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