To study the inhibitory effect of siRNA on cyclin D1 expression and cell proliferation in breast cancer MCF-7 cell line. The siRNA targeting cyclin D1 was chemically synthesized and transfected into MCF-7 cells by oligofectamine. The expression of cyclin D1 was analyzed by quantitive PCR and Western blot, and the cell growth inhibition was measured with CCK-8 assay. Then, cell cycle of the transfected cells was examined by flow cytometry, and cell colony forming ability was measured by soft-agar colony formation assay. After MCF-7 cells were transfected with 10, 50, 100nmol/L siRNA,the expression of cyclin D1 mRNA was respectively suppressed with inhibition rates of 57.85%, 63.22% and 68.02%, and the protein expression was suppressed with inhibition rates of 51.13%, 62.09% and 77.68% respectively. The proliferation of MCF-7 cells was inhibited after transfection with siRNA-cyclin D1, which caused cell cycle arrest at G1 phase and showed less colony forming ability in the breast cancer cell line MCF-7. These results indicate that siRNA-cyclin D1 could be a powerful anti-proliferative tool in breast cancer gene therapy.