Breast cancer resistance to therapy can result from expression of antiapoptotic genes. Survivin is an antiapoptotic gene that is over expressed in most human tumors. RNA interference using short interfering RNA (siRNA) can be used to specifically inhibit survivin expression. A novel siRNA targeting survivin was used to process MCF-7 cells. Cellular survivin mRNA and protein levels were determined by real-time qRT-PCR and Western blot, respectively. Cellular morphology and cell cycle were determined by fluorescence microscopy and flow cytometry. Cell proliferation was measured by MTT assay. Our data showed that the novel survivin-targeted siRNA could efficiently knockdown the expression of survivin, inhibit cell proliferation and cell cycle, especially at the G2/M checkpoint. These data suggest that the siRNA has potential for therapeutic applications.