AU - Nikyar, Arash AU - Bolhassani, Azam AU - Rouhollah, Fatemeh AU - Heshmati, Masoumeh TI - Construction of a Prokaryotic Expression Vector harboring Two HIV-1 Accessory Genes PT - JOURNAL ARTICLE TA - mljgoums JN - mljgoums VO - 15 VI - 2 IP - 2 4099 - http://mlj.goums.ac.ir/article-1-1356-en.html 4100 - http://mlj.goums.ac.ir/article-1-1356-en.pdf SO - mljgoums 2 ABĀ  - Background and objectives: HIV-1 Nef and Vpr antigens have been described as suitable candidates for therapeutic HIV vaccine development. The aim of this study was to generate Nef-Vpr fusion gene construct and to clone the construct into pET-23a (+), a prokaryotic expression vector. Methods: HIV-1 Nef and Vpr genes were PCR-amplified from the pNL4-3 plasmid using specific primers and Pfu DNA polymerase. Results of PCR amplification were visualized by electrophoresis on 0.8% agarose gel. At first, the amplified Nef fragment was cloned into NheI and BamHI restriction sites of pET-23a expression vector. Next, cloning of Vpr gene was performed into BamHI and HindIII restriction sites of the pET-23a-Nef vector. Finally, purity of the recombinant pET-23-Nef-Vpr construct was determined by NanoDrop spectrophotometry. Results: PCR amplification of Nef and Vpr genes was confirmed by detection of ~ 620 bp and ~ 291 bp bands, respectively. Cloning of the Nef-Vpr construct into the vector was confirmed by detection of a ~ 911 bp fragment following enzymatic digestion with NheI and HindIII and sequencing. Conclusion: The successful construction of recombinant fusion plasmid encoding a chimeric Nef-Vpr gene was performed in a prokaryotic expression vector for development of HIV-1 recombinant protein vaccine in near future. CP - IRAN IN - Department of Hepatitis and AIDs, Pasteur Institute of Iran, Tehran, Iran LG - eng PB - mljgoums PG - 11 PT - Original Paper YR - 2021