Objective    To construct the adenoviral recombinant plasmid encoding the genes associated with retinoid-IFN-induced mortality 19 (GRIM19)

and observe its cytotoxic effect on the viral packaging cells and evaluate the expression of GRIM19 in human glioma cell line CHG-5.     Methods    The GRIM19 gene and IRES2EGFP gene were respectively subcloned into pAdTrack-CMV plasmid. After confirmed by restriction enzyme digestion and sequencin

g

the recombinant plasmid of pAdTrack-CMV-GRIM19 was linearized with PmeⅠ and transformed into competent BJ5183 cells containing pAdeasy-1 vector. Then the positive clones with homo-logous recombination (pAd-GRIM19) was confirmed by PacⅠ digestion and transfected into the packaging 293 T cells to generate virus and the viral supernatant containing the GRIM19 gene was confirmed by PCR. After the virus was tittered by using TCID

50

method

CHG-5 cells were infected with the recombinant adenovirus and the expression of GRIM19 was detected by Western blot analysis.     Results    The recombinant adenoviral plasmid pAd-GRIM19 was successfully constructed. The adenoviral titers for GRIM19 and EGFP control recombinant were 5×10

10

U/ml and 8×10

10

U/ml respectively. The recombinant virus infected CHG-5 cells with great efficiency.     Conclusion    The recombinant adenoviral plasmid of pAd-GRIM19 and pAd-EGFP control are constructed successfully. GRIM19 is overexpressed in CHG-5 cells

providing a basis for further study of GRIM19 in vitro and in vivo.