Inhibition of survivin gene expression in EJ cells by RNAi

Objective:To construct eukaryotic expression vectors with the pSilencer2.0 vector for inhibiting human survivin gene by RNA interference,and to detect the effect of the silenced survivin gene on EJ cells.Methods:Two target gene segments were synthesized and cloned into the pSilencer2.0 vector respectively to construct two recombinant eukaryotic expression vectors,pSilencer2.0-SVV1and pSilencer2.0-SVV2,which were identified by enzyme digestion analysis and DNA sequencing.Then the EJ cells were transfected with the recombinant vectors by lipofection and the interference effect was detected by RT-PCR and Western-blot.Results:Enzyme digestion analysis and DNA sequencing showed that two target segments were cloned into pSilencer2.0 vectors correctly.The results of RT-PCR and Western blot indicated that pSilencer2.0-SVV1 and pSilencer2.0-SVV2 vectors could significantly knock down the transcription and expression of survivin gene(P0.01),and the latter was better.Conclusion:pSilencer2.0-SVV1 and pSilencer2.0-SVV2 vectors for survivin gene were constructed successfully.The transcription and expression of survivin gene were inhibited effectively by the constructed vectors in the bladder cancer cell line EJ and the latter was better.This paves way for the subsequent studies.