Construction and identification of lentiviral vector of RNA interference of CDC25B3 gene

Objective To construct a lentiviral vector of RNA interference(RNAi) of CDC25B3 gene.Methods The complementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed,synthesized and cloned into the pGCL-GFP vector,which contained U6 promoter and green fluorescent protein(GFP).The resulting lentiviral vector containing CDC25B3 shRNA was named LV-sh CDC25B3,which was confirmed by PCR and sequencing.293T cells were cotransfected with lentiviral vector LV-shCDC25B3,pHelper 1.0 and pHelper 2.0.All virus stocks were produced by Lipofectamine 2000-mediated transfection.The titer of virus was tested according to the expression level of GFP.Results PCR and DNA sequencing demonstrated that the lentivirus RNAi vector of CDC25B3(LV-shCDC25B3) producing CDC25B3 shRNA was constructed successfully.The titer of concentrated virus was 1×108TU/ml.Conclusion The lentivirus RNAi vector of CDC25B3 has been constructed successfully.