Influence of RNA interference lentiviral vector in E2F-1 on COX-2 in Tca8113 cells

Objective To investigate the influence of RNA interference lentiviral vector in E2F-1 on COX-2 in Tca8113 cells.Methods DNA oligonucleotides targeting gene E2F-1 on appropriate site and one pair of negative control oligonucleotide sequence were synthesized and inserted into pLKO.1-PURO-SP6(Age I/EcoR I) vector.The sequences of two plasmids were identified by DNA sequencer and restriction endonuclease digestion.These recombinant plasmids were transfected into the 293T cells along with lentiviral packing mix by lipofectamine 2000 for the package of lentiviral particles.Then the lentiviral vector particles were transfected into Tca8113 cells and E2F-1 expression in the transfected cells was assayed with Real-Time PCR.The expression of E2F-1 and COX-2 protein was examined by Western-blot.Results It was verified that the specific DNA oligonucleotide was cloned into the vector successfully and the expression of E2F-1 mRNA in Tca8113 cells was significantly reduced after the recombinant plasmids were transfected,compared to the control group.(P0.05).The expression of E2F-1 and COX-2 protein was significantly reduced after transfection.Conclusions E2F-1 gene silencing suppresses the expression of COX-2,indicating COX-2 is the target gene of E2F-1.