Construction and Expression of Human Papillomavirus Type 11 E7 Gene with Recombinant Adenovirus Vectors in Eukaryotic Cells / 中华皮肤科杂志

ABSTRACT

Objective To construct and express human papillomavirus type 11(HPV11) E7 gene with recombinant adenovirus vectors. Methods HPV11 E7 gene was amplified by PCR and directionally cloned into vector pENTR-TOPO to form TOPO-E7 plasmid. E7 gene was transferred into the pAD/CMV/V5-DESTTM gateway vector by LR recombination reaction with pAD/CMV/V5-DESTTM gateway vectors and TOPO-E7 plasmid. The recombination vector was digested by Pac I enzyme and transfected into 293A cell by Lipofectamine method to obtain recombinant adenovirus vectors pAD-E7. Expression of E7 on HaCaT cells infected with pAD-E7 vectors was analyzed by confocal microscopy. Results The recombinant plasmid TOPO-E7 was identified and confirmed with enzyme digestion and sequencing. Recombinant adenovirus vectors pAD-E7 were generated efficiently with a titer of 1.4 ? 107 pfu/mL in transfected 293A cells. E7 protein could be identified in HaCaT cells with confocal microscope 48 h after infected with recombinant adenovirus vector. Conclusions The results indicate efficient expression of HPV11 E7 gene in eukaryotic cells by recombinant adenovirus mediated transfer, which facilitates further research of its function.