Abstract: Objective：To elucidate the role of cell-permeable TAT-N24 fusion peptide inhibiting proliferation of colon cancer cells. Methods ：Detect the cell cycle progression by flow cytometer, analyze the DNA synthesis by BrdU/PI method and certify the roles of TAT-N24 in the tumor. Results ：Western blot shows that TAT-N24 can enter HT29 cells. With the treatment of TAT-N24，the Rb phosphorylation is decreased.In the control group，the number of BrdU positive cells is (52.2±1.88)%, while the TAT-N24 group is (29.9±2.14)%.In control group, cell cycle analysis shows that the number of cells in G0/G1 phases is(55.27± 2.48)%, in S and G2/M phases cells are (26.97±0.94)% and (17.76±1.77)%, but in TAT-N24 group, the number of cells in G0/G1 phases is(65.10±1.79)%, the S and G2/M phases cells are (18.49±0.68)% and (16.40±1.51)%. Conclusion ：TAT-N24 fusion peptide can inhibit the Rb protein phosphorylation, induce the cell cycle arrest and inhibit the DNA synthesis in colon cancer cells.