[Objective] In order to lay the foundation for the further development and utilization of biocontrol strains with good colonization ability, bacterial strains with antifungal activity were isolated, screened and identified from soil. [Methods] Antagonistic experiment was used to screen the specific bacteria against pathogenic fungi and evaluate its inhibitory ability. The strain was classified and identified by morphological characteristics, physiological and biochemical characteristics, sequencing of 16S rRNA gene and analysis of sequences similarity in GenBank. Protease activity of the test strain was measured by Folin-Ciocalteu method. [Results] Brevibacillus laterosporus, preservation number AMCC 100017, was isolated from various kinds of?soil in Tai’an, Shandong province. Plate confrontation experiment indicated that the strain has strong antagonistic effect against various plant pathogenic fungi, especially Fusarium. In addition, it was also verified that the strain could produce highly reactive extracellular protease. [Conclusion] Brevibacillus laterosporus AMCC 100017 is a biocontrol strain wih the development and utilization potential against fungal diseases of crops and fruit trees, as well as the biological control of nematodes.