Construction of pEGFP-E1A eukaryotic expression vector and its inhihition effect to SMMC-7721 hepatoma cell lines in vitro

Objective To construction of recombinant eukaryotic expression vector pEGFP-E1A containing EGFP and Ad5.EIA,in vitro study of inhibition effect to SMMC-7721.Methods The E1A gene was digested from pUC119-E1A plasmid and cloned into pEGFP-C1,a eukaryotic expression vector containing EGFP and neo as selection marker,to form pEGFP-E1A.The pEGFP-E1A was transfected into SMMC-7721 hepatoma cell lines with lipofectin by identification with RT-PCR and immunofluorescence assay.Apoptosis was evaluated using DNA gel electrophoresis.Results Enzyme restriction and sequencing data indicated that the recombinant vector was constructed exactly,RT-PCR and immunofluorescence assay result shown that the pUC119-E1A recombinant eukaryotic expression vector can efficiently express E1A in SMMC-7721 cells,and apoptosis of SMMC-7721 appears at 6 hours.Conclusion The results preliminarily confirmed E1A gene can induced apoptosis SMMC-7721 human hepatoma cell in vitro,which offered the basic experimental conditions for exploring the antitumor activity led by the Ad5.EIA gene.