Construction of Replication Deficient Recombinant Adenovirus of NP9 Protein

Objective:To construct replication deficient recombinant adenovirus vector of human NP9 protein by homologous recombination in bacteria using AdEasy system,and produce adenoviral particles.Methods: The CDS of NP9 gene was obtained from pMD18T-NP9 plasmid by restriction endonuclease digestion, and was inserted into the downstream of CMV promoter of a shutter plasmid pAdTrack-CMV. The recombinant transfer plasmid pAdTrack-CMV-MCP-1 was linearized by Pme I and co-transformed with backbone vector pAdEasy-1 into bacteria BJ5183 for homologous recombination to obtain the recombinant adenoviral plasmid. The recombinant adenoviral plasmid was linearized and was then transfected into HEK293 package cells to produce virus particles. The detection of recombinant adenovirus was characterized by using PCR.Results:The recombinant adenoviral plasmid was successfully established and confirmed by restriction endonuclease digestion. GFP expression was observed on the 5th day after transfection. Recombinant adenovirus was identified by PCR.Conclusion:The achievement of recombinant adenoviral plasmid and recombinant adenovirus of NP9 laid a foundation for further investigation of its function and application.