Objective    To construct the enhanced green fluorescent protein (EGFP) and Rb94 co-expression vector and identify its expression in cultured eukaryocytes.     Methods    PCR was performed to amplify Rb94 gene fragments from cDNA-Rb plasmids. Rb94 gene fragments

digested with BamHⅠand SalⅠ

were harvested and inserted into pEGFP-C1 to construct the pEGFP-C1/Rb94 expression vector. Recombinant plasmid pEGFP-C1/Rb94 was transfected into human retinoblastoma (HXO-Rb44) cells with liposome. Its expression was observed under a fluorescence microscope. Expression of Rb protein was detected by Western blotting. Apoptotic rate of HXO-Rb44 cells was detected by flow cytometry.      Results    The sequence of cloned DNA fragments was fully consistent with that of the Rb94 gene. The Rb94 gene was inserted into the eukaryotic expression vector

pEGFP-C1. The recombinant expression plasmid was successfully transfected into HXO-Rb44 cells. Fluorescent microscopy showed that fluorescence occurred in HXO-Rb44 cells. Western blotting demonstrated the expression of Rb protein. Flow cytometry revealed that the apoptotic rate of HXO-Rb44 cells containing the Rb94 gene was significantly higher than that of other cells.     Conclusion    A recombinant eukaryotic co-expression vector of EGFP and Rb94 can be successfully constructed

which is effectively expressed in HXO-Rb44 cells. The Rb94 gene can effectively inhibit the growth of HXO-Rb44 cells.