Baculoviral expression of recombinant s1 domain of the porcine epidemic diarrhea virus spike (pedv-s1) protein of highly pathogenic pedv in insect cells

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Author: 
Nguyen Van Noi, Yao-Chi Chung and Ta-Chih Cheng
Abstract: 

Background: Highly pathogenic Porcine epidemic diarrhea virus (PEDV) is causing damages to the swine industry and is responsible for loss of economics. Vaccination is the most effective method to prevent and control PEDV infections.
Methods: Recombinant virus-like particles (VLPs) by baculovirus expression vector system have been suggested as a promising platform for new viral vaccines.
Results: In this study we constructed a recombinant baculovirus that was potent to express PEDV-S1 protein; PEDV-S1 protein (S1) which is essential to generate immunogenic VLPs in insect cells. A triplet cassette providing simultaneous and independent expression of S1 gene of PED virus was designed and subjected to synthesis. The cassette was then cloned into pFastBacHTb plasmid and then transformed in to competent Escherichia coli DH10Bac cells and the recombinant bacmids were produced following the homologous Tn7 site-specific transposition. This construction was verified by polymerase chain reaction (PCR) and then transfected into Sf9 insect cells to package new recombinant baculovirus expressing complex proteins of the interest. Restriction map of pFastBacHTb_PEDV-S1 plasmid confirmed the fidelity of the clone. The PCR carried out on the recombinant bacmids as template indicated that a proper homologous recombination has occurred between pFastBacHTb_PEDV-S1 donor plasmid and the bacmid. Following the transfection of new recombinant bacmids to Sf9 cells, cytopathic effects were observed which indicating the successful packaging of recombinant baculovirus. 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 work has proper characteristics to produce PED virus-like particles in Sf9 cells

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