Abstract: In order to screen out the antagonistic bacteria with a good inhibitory effect on grape gray mold,from the root soil of wine grapes,Botrytis cinerea was used as a target strain and the antagonistic bacteria was treated with Oxford cup method. The antagonistic bacterias were identified by morphological,physiological,biochemical characteristics and bacterial 16S rDNA.The antagonistic bacteria growth characteristics and the inhibitory effects of antagonistic bacteria on pathogens were analyzed. The results showed that the T3 strain was screened to have significant inhibition effect on Botrytis cinerea.T3 was identified as Bacillus subtilis and entered a stable phase after being cultured for 24~48 h. The optimal growth temperature of T3 strain was 37 ℃ and its optimal pH was 7.0.The MIC of T3 was 3.6×10⁵ CFU/mL and the MBC was 3.6×10⁶ CFU/mL. The 3.6×10⁹ CFU/mL of T3 strain had a bacteriostatic diameter of 31.6±0.2 mm(P<0.05)against Botrytis cinerea and its bacteriostatic effect price was 158.0±1.0 mm/mL(P<0.05). When observed under the scanning electron microscope,Botrytis cinerea showed mycelium deformity,twisting,apical swelling,protoplast contraction and spores sparsed.T3 strain inhibited mycelial growth and spore germination. It provided the new microbial resource for the biological control of wine grape gray mold and had further research value.