Construction and expression of recombinant adenovirus pAdxsi-GFP-HIF-KGF containing hypoxia inducible factor gene and keratinocyte growth factor gene

Objective: To construct a recombinant adenovirus(pAdxsi-GFP-HIF-KGF) encoding human hypoxia inducible factor 1α gene(HIF-1α) and human keratinocyte growth factor gene,and investigate its expression in lung epithelium cells. Methods: HIF-1α gene from A549 in hypoxia condition and KGF gene from plasmid pIRES2-EGFPKGF were subcloned into shuttle vector pShuttleCMVEGFP. A shuttle vector pShuttle-GFP-HIF-KGF was obtained. Then it was cotransformed into adenoviral backbone plasmid pAdxsi to make homologous recombination with HIF-1α gene and KGF gene. The recombinant adenovirus was infected with A549,and the expression level of HIF-1α protein and KGF protein were evaluated by ELISA. Results: The recombinant adenovirus vector for HIF-1α gene and KGF gene(pAdxsi-GFP-HIF-KGF) was successfully constructed. The amount of HIF-1 protein and KGF protein was(56. 36 ± 4. 53) ng / ml and(60. 20 ± 2. 92) ng / ml in supernatant at 48 h after transfection,respectively. Conclusion: A recombinant adenovirus pAdxsi-GFP-HIF-KGF,encoding human hypoxia inducible factor 1α gene and keratinocyte growth factor gene,was constructed and expressed successfully in A549 cells. It has further application prospect for the prevention and treatment of lung injury.