Abstract: Objective:To evaluate the inhibitory effect of RNA interference (RNAi) on the expression of epithelial cell adhesion molecule (EP-CAM) mRNA and protein in human gallbladder cancer cells.Methods:Four recombinant plasmids targeting human EP-CAM gene were transfected into GBC-SD cells by Lipofectamine 2000 after DNA sequencing;RT-PCR and Western blotting were used to detect the mRNA and protein expression of EP-CAM.Results:Four EP-CAM-targeted microRNA (miRNA) -1,-2,-3and-4 were successfully inserted into the plasmid vector,and the coding sequences of the obtained miRNA were consistent with the designed fragments.The results indicated that four recombinants plasmid could effectively knock down the expression of EP-CAM mRNA and protein in human gallbladder cancer cells.Conclusions:The recombinant plasmid pcDNA^TM6.2-GW/EmGFPmiR miRNA targeting human EP-CAM gene can effectively down-regulate the expression of EPCAM mRNA and protein in human GBC-SD cells.