Abstract: Interference sequences of different positions of CSFV NS3 gene were designed,these sequences were annealed and cloned to interferece plasmid. Recombinant interferencing plasmids were named pGene-NS3-1, pGene-NS3-2, pGene-NS3-3 and pGene-NS3-Negative (negative control) by restriction analyzing and sequencing. These recombinant plasmids transfected PK-15 cells and G418 screened. CSFV were inoculated in cells,which cultured largely.Collected cells were analyzed for Real-time PCR and ELISA. The results of Real-time PCR indicated that mRNAs were silenced by pGene-NS3-1,pGene-NS3-2 and pGene-NS3-3 and inhibition ratio are 63%、46%、49%.The results of ELISA indicated that CSFV’s propagation were inhibited in PK-15 cells by pGene-NS3-1,pGene-NS3-2 and pGene-NS3-3 differently.