Abstract: Root rot represents a significant disease affecting the cultivation of Platycodon grandiflorum. The screening of biocontrol strains with antagonistic properties against this disease can provide valuable microbial resources for the environmentally friendly prevention and control of Platycodon grandiflorum diseases. In this investigation, high-throughput bacterial isolation techniques were utilized to isolate and purify endophytic bacteria from the roots of Platycodon grandiflorum. An antagonistic bacterial strain R34B7, was identified through the plate confrontation culture method, exhibiting a notable inhibitory rate of 52.18% against the pathogen causing root rot. Morphological characteristics, physiological and biochemical properties, and molecular identification results collectively confirmed that the antagonistic endophytic bacterium R34B7 belonged to the species Serratia plymuthica. The control efficacy of strain R34B7 against Platycodon grandiflorum root rot was assessed using tissue-cultured seedlings of Platycodon grandiflorum, revealing a disease control efficacy of 44.44%. Furthermore, the fermented supernatant of strain R34B7 demonstrated considerable inhibitory effects on both the mycelial growth and spore germination of the pathogen. By examining the extracellular hydrolytic enzymes and growth-promoting factors of strain R34B7, it was discovered that this strain possesses the abilities to produce protease, chitinase, fix nitrogen, solubilize phosphorus, synthesize siderophores, and produce indole-3-acetic acid, indicating its potential for growth promotion. The antagonistic bacterium R34B7 identified in this study exhibits promising biocontrol activity against root rot and holds considerable potential for further development and utilization.