Objective      To construct a recombinant adenovirus vector of rat erythropoietin (EPO) and observe its expression in rat spiral ganglion neurons (SGNs)．        Methods      EPO gene fragment was amplified and subcloned into shuttle plasmid pAdTrack-CMV-EPO. The constructed recombinant shuttle plasmid pAdTrack-CMV-EPO was identified by restriction analysis

PCR and sequencing

then linearilized with PacⅠ and transformed to competent AdEasy systems. The obtained rec

ombinant adenovirus vector AdEPO was digested with PacⅠ and transfected to 293 cells for packaging

and the prepared recombinant adenovirus was amplified for titration and identification by RT-PCR and Western blotting. The vector had been transfected into the cultured SGNs

and EPO expressions were detected by immunofluorescence and Western blot analysis respectively.       Results      After the incision enzymes of KpnⅠ

HindⅢ and sequencing

pAdTrack-CMV-EPO was accomplished. Recombinant adenovirus AdEPO was successfully packaged in 293 cells with a titer of 1.17×10

11

IU/ml after amplification，and successfully expressed in 293 cells. Both recombinant shuttle plasmid and recombinant adenovirus vector AdEPO were constructed correctly. EPO level was increased obviously after the virus transfection in SGNs either with immunofluorescence or Western blot assays.       Conclusion      The recombinant adenovirus vector of EPO is successfully constructed and significantly enhances the EPO levels after its transfection into SGNs.