Abstract: To clarify the mechanism of resistance of Bacillus velezensis TP-1 to gray mold, a marker strain TP-1^R was screened for resistance to 300 μg/mL of rifampicin by an antibiotic marker method in this study. The genetic stability of the resistance of the marker strains and their antagonism to gray mold of grapes were studied, and their colonization in grapes and their effect on the activity of defence enzymes in the grapes were analysed. The results showed that the labeled strain could still grow stably in the medium containing rifampicin after 15 times of subculture, and the antagonistic effect of the labeled strain against gray mold was not significantly different from that of the original strain. The colonization amount of the labeled strain reached the peak of 4.32×10⁶ CFU/g on the 15th day of storage (20 ℃), and was still 3.11×10⁶ CFU/g on the 30th day of storage, indicating that strain TP-1 could be stably colonized on grapes. Inoculation of strain TP-1 fermentation broth could effectively inhibit the occurrence of gray mold and resultantly reduced the decay rate of grapes. During storage, the activity of grape defence enzymes PAL, PPO and APX increased first and then decreased, and the activity of the three enzymes in the group treated with the antagonistic bacterium TP-1 was significantly higher than CK (P<0.05). At the 15th day, the activities of PAL, PPO and APX in the antagonistic bacterium treatment were 1.23, 1.19 and 2.01 times higher than CK. The colonization of strain TP-1 on grape could enhance activities of a set of defense-related enzymes, including PAL, PPO and APX, and enhance the resistance of grape to gray mold to a certain extent. This study provided a scientific basis for revealing the mechanism involved in the biocontrol effectiveness of Bacillus velezensis TP-1.