Purpose Ovarian cancer is a devastating gynecological disease which is considered the major cause of cancer fatality around the world. The down-regulation of microRNA-33b (miR-33b) was reported in some malignancies. Hence, we transfected the miR-33b mimic into SKOV3 cells and evaluated the impacts of this interference on the growth and migration repression of these tumor cells as well as on targeted genes expression. Methods In our study, transfecting the miR-33b mimic and inhibitor, negative control (NC), and NC inhibitor were established using Lipofectamine 2000. The cytotoxic effects of miR-33b were evaluated by MTT. To assess the miR-33b effects on cell migration, a scratching test was applied. The expression levels of miR-33b, ADAMTS, C-Myc, MMP9, K-Ras, and CXCR4 were evaluated using qRT-PCR. Results These findings indicate that transfection of miR-143 mimic had no marked effects on the SKOV3 cell line. As expected, miR-33b relative expression levels were as follows: miR-33b mimic >NC and NC inhibitor >miR-33b inhibitor (p < 0.01). Moreover, transfected miR-33b mimic could suppress SKOV3 cells' proliferation, whereas transfected miR-33b inhibitor could promote cell proliferation (p < 0.01). MiR-33b overexpression significantly down-regulated the MMP9, CXCR-4, c-Myc, ADAMTS, and K-Ras mRNA levels (p < 0.05). Conclusion As expected, these results confirm the tumor-suppressive effect of miR-33b in the SKOV3 ovarian cancer cell line by reducing cell survival, proliferation, and migration.