Construction and Identification of Recombinant Adenovirus Vector Containing NY-ESO-1 Gene

Objective To construct recombinant advenovirus plasmid containing human tumor-specific antigen NY-ESO-1 gene by using the method of homologous recombination in bacteria.Methods The NY-ESO-1 cDNA primers were designed,the DNA sequence of NY-ESO-1 was amplified from recombinant vector pET15b-NY-ESO-1 and ligated into pShuttle-IRES-hrGFP-2.The recombinant plasmid was named after pShuttle-NY-ESO-1-EGFP and was identified with PCR and EcoR Ⅴdigestion.pShuttle-NY-ESO-1-EGFP was linealized with PmeⅠand transformed into ultracompletent BJ5183 containing pAdeasy-1,then recombinant advenovirus pAd-NY-ESO-1-EGFP was constructed by homologous recombination in bacteria.The recombinant adenovira1 plasmid pAd-NY-ESO-1-EGFP was identified by PacⅠdigestion and DNA sequencing.Results There were two bands,4.5kb and larger than 23 kb when pAd-NY-ESO-1-EGFP was digested with PacⅠ.A 340bp NY-ESO-1cDNA fragment was amplified by PCR.Conclusion The recombinant adenoviral plasmid containing NY-ESO-1 was successfully constructed with homologous recombination in bacteria.This study provides a basis for the preparation of recombinant adenovirus expressing NY-ESO-1 and tumor-specific antigen NY-ESO-1 DNA vaccine for research in cancer therapy.