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Background: Genetic variability of influenza viruses causes new epidemics worldwide annually. Development of a new vaccine for prophylaxis of influenza virus has been amajor objective in recent years. The aim of this study was to construct a recombinant baculoviruscapable of expressing the two surficial antigenic glycoproteins, hemagglutininand neuraminidase, as well as matrix proteinsof swine influenza (H1N1) simultaneously and independently. Materials and Methods: In this experimental study, first, a triplet cassette providing simultaneous and independent expression of target proteins was designed and subjected to synthesis. It was then cloned into pFastBac1 donor plasmid. Competent E.ColiDH10Bac cells were transformed by donor clone and the recombinant bacmids were produced following homologous transposition. This construction was verified by PCR and then transfected into Sf9 insect cells to package new recombinant baculoviruses. Results: Restriction map of pFastBacI HNM1 donor plasmid confirmed the fidelity of the clone. The results of PCR done on the recombinant bacmidas template indicated that a proper homologous recombination has occurred between pFastBacI HNM1 donor plasmid and the bacmid in E.ColiDH10Bac host cells. Protein analysis of the infected Sf9 cells showed that all target proteins were efficiently expressed at the same time. Conclusion: The recombinant baculovirus constructed in this studypossesses proper characteristics to produce swine influenza virus-like particles in Sf9 cells.

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Background: Influenza type A virus is one of the most important viral agents in human respiratory diseases. The genetic variability of the influenza viruses leads to the incidence of new epidemics worldwide. Hence, there is a growing need for rapid and effective new methods capable of detection and differentiation of influenza virus circulating strains. This study was done to develop a method for rapid differentiation of the subtypes of influenza type A virus. Materials and Methods: In this experimental study, reverse-transcription and polymerase chain reaction (RT-PCR) were performed using a primer set based on M gene of H1N1, H3N2, H5N1, and H9N2 influenza subtypes. Then the amplified fragments were subjected to digestion using subtype specific restriction endonuclease enzymes. Results: The results of PCR reaction showed that the primer pair of the M gene was specific and capable of amplifying all influenza subtypes understudy. Also, different restriction fragment length polymorphism patterns (RFLP) were generated using enzyme digestion reaction on the amplified segment of M gene. Conclusion: RT-PCR and RFLP analysis of the M gene can be employed as a useful method for differentiating influenza virus subtypes

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Background: The importance of VP2 protein of canine parvovirus to bind to human cancer cells and to detect the virus in veterinary detection kits has motivated a lot of research on the production of this protein. In this project, a surface gene of canine parvovirus (VP2) was cloned and expressed in a prokaryotic vector system and its expression was optimized in a specific cell-free prokaryotic expression system. Materials and Methods: In this experimental study, plasmid pET-21aVP2 was constructed by cloning the PCR product of VP2 gene of canine parvovirus into the plasmid expression pET-21a vector. The best sequence was analyzed through PCR and it was followed by confirmation with sequencing and restriction digestion. To produce VP2 protein, plasmid pET-21aVP2 was transferred into Escherichia coli, Rosetta (DE3) strain, and the expression of this protein was induced by IPTG. The production of VP2 protein in both systems was evaluated using SDS PAGE technique. The expressed protein was checked with monoclonal antibody against VP2 protein by Western blotting technique. Results: Successful cloning of VP2 protein was confirmed by enzymatic digestion and sequencing. The expression of VP2 protein in bacterial and cell-free prokaryotic systems was verified by SDS PAGE and the specific band in Western blotting also confirmed the VP2 protein. Conclusion: The results of this study showed that VP2 gene was amplified in the cloning phases and it was successfully cloned in the expression vector. Protein expression was confirmed in both bacterial and cell-free prokaryotic systems.