Objective: To investigate microRNA-25 (miR-25) expression in rectal carcinoma cells and its action. Methods: The miR-25 expressions in several different rectal cell lines were determined, and then the changes in biological behaviors in rectal cancer cells after up- or down-regulation of miR-25 expression with miR-25 precursor (pre-miR-25) or miR-25 inhibitor (anti-miR-25) transfection were examined. Results: Compared with normal rectal mucosal tissue, the miR-25 expressions in all tested rectal cancer cells were significantly increased with varying degrees (all P<0.05). In rectal cancer HR-834 cells with relatively low miR-25 expression after pre-miR-25 transfection and in rectal cancer SW-837 cells with high miR-25 expression after anti-miR-25 transfection, the proliferation, cell cycle and apoptosis showed no significant change (all P<0.05), but the invasion and migration abilities in HR-834 cells were increased, and SW-837 cells were decreased significantly (all P<0.05). Conclusion: The miR-25 expression is elevated in rectal cancer cells, and its degree of increase is closely related to the invasion and migration abilities of rectal cancer cells.