Construction of eukaryotic expression vector of EBNA2 and functional detection of its product

    WANG Yaoª²Chun£¬ZHANG Ping£¬LI Wei£¬HAN Hua

    Department of Medical Genetics and Development Biology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To construct the eukaryotic expression vector of EBNA2 and to express it in eukaryotic cells and detect the function of its product. METHODS: EBNA2 gene was obtained by PCR and DNA recombination from the plasmid pSG5ª²EBNA2. PCR product of EBNA2b was then inserted into plasmid pMDª²18T and sequenced. The full length EBNA2 gene was cut and inserted into eukaryotic expression plasmid pCMVª²HA. Using Lipofectaª²mineTM 2000, the plasmid pCMVª²EBNA2 was transfected into Cosª²7 and then detected by Western blot and immunofluorescence. The activity of EBNA2 was detected by luciferase reporter assay. RESULTS: The EBNA2 gene was successfully cloned and its sequence was identical to that in GenBank. After plasmid pCMVª²EBNA2 was transfected into Cosª²7, the expressed product of EBNA2 was detected by Western blot and immunofluorescence. The transcriptional activity of EBNA2 was demonstrated by reporter assay. CONCLUSION: The eukaryotic expression vector pCMVª²EBNA2 has been successfully constructed and successfully expressed in eukaryotic cell line. The EBNA2 protein has transcriptional activation. ......