Avicenna Journal of Medical Biotechnology، جلد ۱۳، شماره ۳، صفحات ۱۳۱-۱۳۵
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عنوان انگلیسی Expression and Characterization of Two DNA Constructs Derived from HIV-1-vif in Escherichia coli and Mammalian Cells
چکیده انگلیسی مقاله Background: Acquired immunodeficiency syndrome (HIV/AIDS) is still a major global concern and no effective therapeutic vaccine has been produced to prevent the problem. Among HIV-1 proteins, vif as a basic cytoplasmic protein of HIV-1 is involved in late stages of viral generation and plays important role in HIV-1 virion replication. It also increases the stability of virion cores, which probably inhibits early degradation of viral entry. Therefore, it seems rational to apply this protein as a vaccine based on its impact on HIV-1 life cycle. This study aimed at cloning, expression and production of vif protein as an HIV-1 vaccine candidate. Methods: In this study, vif sequence was amplified from pLN4-3 plasmid including HIV-1 vif gene and then cloned in pET23a to generate the recombinant plasmids of pET23a/vif with hexahistidine tags. BL21 competent cells were transformed to obtain the protein of interest. Ni-NTA column was used to purify the protein of interest and western blotting confirmed vif protein using anti-His tag antibody. In order to express the gene of interest in eukaryotic cells, vif was sub-cloned into pEGFP plasmids and HEK 293-T cells were transfected. Flow cytometry was then applied to evaluate GFP expression. Results: vif protein was expressed in BL21)DE3) strain and identified as a23 kDa band in SDS-PAGE and confirmed by anti-His antibody in western blotting. The purified protein concentration was 173.3 μg/ml using Bradford assay. HEK-293T cells were successfully transfected by recombinant pEGFP plasmids and flow cytometry confirmed the cell transfection. Conclusion: vif protein can be expressed in mammalian cells and may be a proper protein subunit vaccine candidate against HIV-1.
کلیدواژه‌های انگلیسی مقاله HIV-1, Vaccines, Vif protein
نویسندگان مقاله | Fatemeh Zamani
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran


| Azam Bolhassani
Department of Hepatitis, AIDS and Blood Borne Diseases, Pasteur Institute of Iran, Tehran, Iran


| Sepideh Shahbazi
Department of Hepatitis, AIDS and Blood Borne Diseases, Pasteur Institute of Iran, Tehran, Iran


| Ahmad Faghih
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran


| Seyed Mehdi Sadat


نشانی اینترنتی https://www.ajmb.org/En/Article.aspx?id=40465
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