Abstract: In order to obtain a Bacillus with biocontrol effects on tobacco brown spot disease, 187 strains isolated from tobacco leaves were used in soaking tobacco seeds, and 6 strains that could promote seed germination were selected by measuring the germination rate. The results from antagonistic, detached leaf inoculation, greenhouse bioassay experiments showed that strains FJ1 and FJ1-6 could promote tobacco growth and had control effects on brown spot disease. Physiological, biochemical and molecular biological identification revealed that FJ1 and FJ1-6 belonged to Bacillus atrophaeus and Bacillus velezensis. The study of Biocontrol characteristics showed that the two strains contained genes involved in the synthesis of lipopeptide antibiotics Surfactin, Fengycin and Iturin, and the crude lipopeptide extracts had a significant inhibitory effect on the growth of the mycelium of brown spot disease, with the inhibition rates of 56.9% and 65.0% respectively. The results show that FJ1 and FJ1-6 have excellent growth-promoting and disease-preventing properties and significant seed germination effects. They can be applied to tobacco seedling cultivation and tobacco brown spot control and provide effective strain resources for the development of biocontrol agents.