Late blight of potato is caused by the pathogen Phytophthora infestans, which has been considered to be the most destructive disease affecting potato crops worldwide. In recent years, the use of antagonistic microorganisms to control potato late blight has become a green and environmentally friendly means of disease control, greatly reducing the use of chemical pesticides. To obtain antagonistic bacteria with a high biocontrol effect against potato late blight, a total of 16 antagonistic bacterial strains with an inhibition rate of more than 50% against P. infestans were screened from potato rhizosphere soil by double-culture method, among which the bacterial isolate (X3-2) had the strongest inhibitory activity against P. infestans, with an inhibition rate of 81.97 ± 4.81%, respectively, and a broad-spectrum inhibitory activity. The bacterial isolate (X3-2) was identified as Bacillus velezensis based on its 16S rDNA gene sequence and morphological as well as biochemical properties. The results of our in vitro experiments demonstrated that X3-2 was a potent inducer of resistance in potato tubers and leaflets against late blight. In greenhouse experiments, it was confirmed that the biological preparation X3-2 exhibits an anti-oomycete effect, demonstrating a significant control efficacy on potato late blight. Further analyses showed that the antagonistic substances of X3-2 were distributed both intracellularly and extracellularly. In addition, screening for plant-growth-promoting (PGP) traits showed that X3-2 has the ability to produce siderophores and secrete indole acetic acid (IAA). The findings from this research suggest that B. velezensis X3-2 exhibits promise as a biocontrol agent for managing late blight. In the future, the composition and mechanism of the action of its antimicrobial substances can be studied in depth, and field trials can be carried out to assess its actual prevention and control effects.