en 27920885 <p>Background: As a drug target and an antigenic agent, HIV-1 protease (HIV-1 PR) is at the center of attention for designing anti-AIDS inhibitors and diagnostic tests. In previous studies, the production of the recombinant protease has been faced with several difficulties; therefore, the aims of this study were the easy production, purification of the soluble form of protease in <em>E. coli</em> and investigation of its immunoreactivity.<br /> Methods: Protease coding region was isolated from the serum of an infected individual, amplified by RT-PCR and cloned into PTZ57R using TA-cloning. Protease coding frame was isolated by PCR and cloned in pET102/D. TOPO expression vector and cloned protease was expressed in <em>Escherichia coli (E. coli)</em> BL21. Produced recombinant protein was purified by affinity Ni-NTA column and protein concentration was checked by BCA protein assay kit. Subsequently, immunoreactivity of recombinant protease (rPR) was assayed by Western blotting and ELISA.<br /> Results: Cloning of the HIV protease by TOPO cloning system in pET102/D.TOPO was confirmed with PCR and sequencing. The concentration range of purified recombinant protein was 85 to 100<em> &mu;g/ml</em>. Immunogenicity of rPR was confirmed by Western blotting and ELISA.<br /> Conclusion: Soluble production of recombinant HIV-1 protease (HIV-1 rPR) was performed successfully. This recombinant protein disclosed 86% specificity and 90% sensitivity in immunoassay tests.</p> Human immunodeficiency virus, Molecular cloning, Protease, Recombinant proteins 175 181 https://www.ajmb.org/En/Article.aspx?id=254 https://www.ajmb.org/PDF/En/FullText/254.pdf AsaadAzarnezhadDepartment of Medical Genetics, Faculty of Medicine, Tehran University of Medical Science, Tehran, Iran1035ZohrehSharifiBlood Transfusion Research Center, Institute for Research and Education in Transfusion Medicine, Tehran, Iran640RahmatollahSeyedabadiDepartment of Molecular Medicine and Genetics, Faculty of Medicine, Hamedan University of Medical Sciences, Hamedan, Iran1036ArshadHosseiniDepartment of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences, Tehran, Iran1037BehroozJohariDepartment of Medical Biotechnology, Faculty of Allied Medicine, Iran University of Medical Sciences, Tehran, Iran1038MahsaSobhani FardStudent Research Center, Hamedan University of Medical Sciences, Hamedan, Iran1039