Recombinant Adenovirus Vector Mediated Expression of HCMV UL49 Gene in Mice / 中国生物工程杂志

ABSTRACT

Human cytomegalovirus (HCMV) is extremely species specific and does not replicate in experimental animal tissues.To overcome the problem and establish suitable animal models for studying antiviral strategies,the expression of HCMV UL49 gene was explored in mice.UL49-GFP gene was subcloned into the adenovirus shuttle plasmid pDC316,the products(pDC316-UL49-GFP)were co-transfected with helper plasmid pBHGloxE1,3Cre into HEK293 cell lines by liposome reagent,recombinant adenovirus(Ad-UL49-GFP) was generated and confirmed by PCR and Western blot.Ad-UL49-GFP was propagated in 293 cells and purified.The titer of viral stocks was determined by end-point dilution assay.The purified adenoviruses were delivered into mice via the tail vein injection.Fluorescence quantitative PCR and Western blot experiments were used to examine the tissue distribution and duration of UL49 gene expression.The results showed that the recombinant adenovirus were present in vivo.The expression level in tissues arranged in descending order was liver,spleen,kidney,heart and lung.3 days after injection,the liver,spleen,kidney,heart and lung expressed protein UL49 in high lever and then declined gradually.14 days after injection,UL49 protein expression was disappear in some organs except liver and spleen.In conclusion,transgene animal model carrying UL49 gene was successfully established.Therefore,the system may be suitable for selecting anti-HCMV drugs targeting UL49 gene.