Objective：To explore the effects of Geminin gene silencing on the biological behavior of human gliomas and the possible mechanism. Methods：Totally 20 pairs of fresh glioma specimens collected from patients who received a surgical resection in the Department of Brain Surgery in the Affiliated Hospital of Nantong University from 2018 to 2019 were analyzed by qRT-PCR combining GEPIA database to study the expression of Geminin gene in glioma tissues. The siRNA sequences that effectively targeted Geminin gene and the negative control sequences were designed and transfected into glioma U251 cells using Lipofectamine- 2000 reagent. The two groups were defined as siRNA-Geminin group and siRNA-NC group，respectively. In 48 h after transfection，qRT-PCR was used to detect the expressions of Geminin mRNA in each group. Then CCK8 method，Transwell assay，and Western blot were used to detect the cell proliferation，migration and invasion ability and the levels of Geminin，HBO1 and Cdt1 protein expression respectively. Results：Results of both qRT-PCR and GEPIA database analyses showed that the expression of Geminin mRNA in glioma tissues was significantly higher than that in adjacent tissue. The siRNA interference in Geminin gene could significantly inhibit its expression in U251 cells. Compared with siRNA-NC group，the cell proliferation ability，cell migrationand cell invasion quantity of the siRNA-Geminin group were significantly reduced，and the expression levels of Geminin，HBO1，and Cdt1 proteins were significantly lowered. Conclusion：Geminin is highly expressed in glioma. Knockdown of Geminin could inhibit the proliferation，migration，and invasion of U251 cells，possibly by down-regulating HBO1 and Cdt1. Thus，Geminin may be expected to be a new molecular target for the treatment of human glioma.