BACKGROUND: The ability of plant growth-promoting rhizobacteria (PGPR) to alleviate iron deficiency-induced chlorosis in plants has been widely reported, but the role of siderophores in the re-greening process has rarely been investigated. In this study, the Priestia megaterium ZS-3 (ZS-3) siderophore was first characterized, and a 100-fold concentration of the crude extract of the siderophore was extracted by solid-phase extraction and used to inoculate Arabidopsis thaliana to investigate whether the ZS-3 siderophore could alleviate plant iron deficiency-induced chlorosis in the presence of an insoluble iron source and to determine how it promoted plant growth. RESULTS: The results indicated that -Fe + Fe CONCLUSIONS: This study revealed that the ZS-3 siderophore Ferrioxamine E [M + Fe-2 H] enhances plant iron uptake and transport activity at the transcriptional level, confirming the important role of the ZS-3 siderophore in plant iron deficiency status, and the results suggest that the ZS-3 siderophore helps plants acquire iron, alleviates plant chlorosis and promotes plant growth through mechanism I of plant iron acquisition. In this study, we closely linked the structural characterization and quantification of siderophores with Fe deficiency-induced chlorosis to elucidate the promotional mechanism of siderophores in Fe-deficient environments.