Abstract: 【Objective】 The objective of this study is to investigate bacterial colonization in rice seedlings by GFP-labeled Bacillus pumilus DX01 and lay a solid foundation for the application of phytopathogenic biocontrol. 【Method】 Genetically stable mutants of B. pumilus DX01 with a strong GFP expression were identified from the GFP-labeled Tn5 insertion library and used for tracing the bacterial colonization in rice seedlings. 【Result】 A total of 1 467 mutants were subjected to qualitative and quantitative analyses of GFP expression by using fluorescence microplate reader, fluorescence activating cell sorter and fluorescent microscopy, and eventually 8 mutants with significantly enhanced GFP expression were identified. Meanwhile, the copy numbers of exogenous DNA integration in the genome of these mutants were detected. The GFP-tagged bacterial cells that released into the rhizosphere of rice seedlings could survive for more than 15 days. Furthermore, decrease of the B. pumilus bacterial cells in undisinfected soils showed more slowly compared with corresponding sterile soils. 【Conclusion】 B. pumilus mainly locates in root hair region and later root branch. Meanwhile, the bacterial film formed on rice root surface was also confirmed. B. pumilus can invade plant through the wound of roots or by young root hairs, and they distribute mostly in root cortex cells and intercellular spaces of cortical layer. The marked bacteria were often found in the vascular bundle of rice seedlings. In conclusion, B. pumilus displayed a good colonization ability in the rhizosphere soils of rice seedlings.

Key words: Oryza sativa , Bacillus pumillus , Tn5 transposition , green fluorescent protein , colonization