Many root-colonizing Pseudomonas spp. exhibiting biocontrol activities produce a wide variety of secondary metabolites with antibiotic activity against plant pathogens in the rhizosphere. Among these metabolites, 2,4-diacetylphloroglucinol (DAPG) is one of the most typical antibiotics in model biocontrol strains of Pseudomonas protegens. We screened DAPG-producing strains among 2,800 fluorescent pseudomonads isolated from the field in Japan. In addition to P. protegens, several other strains close to, but different from, P. protegens have been identified and found to exhibit biocontrol activity. Recent advances in genomic research on these strains revealed that they build up strain-specific genomic repertoires for the biosynthesis of secondary metabolites and niche adaptation. In this mini-review, we introduced our recent genome-based characterization of P. protegens and related strains. Comparative genome analyses in combination with different bioassays have become a powerful tool for the identification of novel biocontrol factors of potentially beneficial strains.