Construction of Recombinant Adenovirus Vector Carrying Human p21~ WAF1 Gene by Homologous Recombination in Bacteria

Objective To construct recombinant adenovirus vector carrying human p21~ WAF1 gene by homologous recombination in bacteria.Methods p21~ WAF1 gene was subcloned into pshuttle-cmv transfer vector,and positive recombinant psh-p21 was linearized with PmeⅠ,then transformed to competent AdBJ5183 cells transformed with adenovirus skeletal vector pAdEasy-1 in advance.The recombinant adenovirus vector pAdp21 was identified by digestion with PacⅠ and BstxⅠ as well as amplification by PCR,then linearized with PacⅠ and transfected to 293 cells to package recombinant adenovirus particles.The recombinant adenovirus was identified by RT-PCR,and its stability was analyzed by PCR.Results A 500 bp fragment was amplified by PCR from the constructed recombinant adenovirus vector pAdp21,which was consistent with that expected.The rAdp21 showed typical character of adenovirus under electron microscope.Conclusion Recombinant adenovirus vector pAdp21 was successfully constructed,and recombinant adenovirus rAdp21 was prepared.