Abstract
Increasing therapeutic applications for recombinant human interferon-gamma (rhIFN-γ), an antiviral pro-inflammatory cytokine, has broadened interest in optimizing methods for its production. We herein describe a unicellular eukaryotic system, Leishmania tarentolae, a Trypanosomatidae protozoan parasite of gecko Tarentola annularis, which has recently been introduced as a candidate for heterologous gene expression. In this study, the hIFN-γ cDNA was amplified from phyto-hemagglutinin-stimulated peripheral blood mononuclear cells of a healthy blood donor using RT–PCR. In order to express, the rhIFN-γ protein, the resulting cDNA was cloned in two expression cassettes (each containing one copy of hIFN-γ cDNA) and integrated into the small subunit of ribosomal RNA gene of L. tarentolae genome by electroporation. Transformed clones were selected in the presence of appropriate antibiotics. Western blotting of rhIFN-γ and ELISA confirmed the expression and production of 9.5 mg of rhIFN-γ protein/l respectively.
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Acknowledgments
The authors thank Dr. Anis Jafari, of Molecular Biology Dept. of Pasteur Institute of Iran for her comments and suggestions. This work was supported by grant No.397 of Pasteur Institute of Iran.
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Davoudi, N., Hemmati, A., Khodayari, Z. et al. Cloning and expression of human IFN-γ in Leishmania tarentolae . World J Microbiol Biotechnol 27, 1893–1899 (2011). https://doi.org/10.1007/s11274-010-0648-4
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DOI: https://doi.org/10.1007/s11274-010-0648-4