Abstract: Objective To explore role of miR-206 in modulating the growth and chemotherapy sensitivity in ovarian cancer cells. Methods Real-time PCR was used to detect the expression of miR-206 in ovarian cancer and normal ovarian tissues. Ovarian cancer SKOV3 cells were transfected with a miR-206 mimic or a specific inhibitor of miR-206, and MTT assay and flow cytometry were used to detect the changes in cell growth and cell cycle transition. Western blotting and luciferase reporter gene assay were employed to identify the target gene and signal pathways of miR-206. The effect of miR-206 on the sensitivity of ovarian cancer cells to 5-Fu was assessed. Results miR-206 was down-regulated in ovarian cancer tissues compared to normal ovarian tissues. Transfection of SKOV3 cells with the miR-206 mimic resulted in obvious growth suppression and delayed cell cycle transition from G1 to S phase by suppressing CDK4, c-Myc, and CCND1 expressions. Transfection with the miR-206 inhibitor obviously promoted the cell growth and significantly increased CDK4 expression in the cells. Luciferase reporter gene assay indicated that miR-206 could directly bind to the 3’UTR of CDK4 gene and reduce the activity of luciferase. Transfection of SKOV3 cells with miR-206 significantly lowered the IC50 of 5-Fu to enhance the chemotherapy sensitivity of the cells to 5-Fu. Conclusion As a potential tumor suppressor, miR-206 directly targets CDK4 to suppress the cell growth and enhance the chemotherapy sensitivity to 5-Fu in ovarian cancer cells in vitro.