Objective: To investigate the effect of the expression level of Rab25 gene on the biological behaviors of breast cancer cells and the relation of its action with Her-2/neu. Methods: The Rab25 gene in different breast cancer cell lines and Her-2/neu gene in different breast cancer cell lines with stable Rab25 gene expression were increased or decreased by using plasmid transfection and RNA interference techniques. The proliferative activity, colony formation and invasive ability in each group of cells were determined. Results: The proliferation activity, colony formation rate, and invasion ability were all significantly increased in groups of cells transfected with Rab25 gene compared with their corresponding groups of cells with Rab25 gene interference, ogrinal cell lines and negative control group of cells (all P<0.05). The proliferation activity, colony formation rate, and invasion ability in different breast cell lines stably expressing Rab25 gene showed no significant alteration regardless of the Her-2/neu gene expression increase or decrease (all P>0.05). Conclusion: Rab25 gene exerts a promoting effect on growth, proliferation and invasiveness in breast cancer cells, and the expression level of Her-2/neu gene can not influence its action.