Abstract: 【Objective】The purpose of the study was to investigate the colonization characteristics of rhizobia in micropropagated sugarcane roots,the effects of inoculation rhizobia on the growth of sugarcane and the nitrogen fixation efficiency in association with sugarcane,and to provide a theoretical basis for the application of rhizobia in sugarcane production.【Method】The pPROBE-pTetr-TT plasmid containing green fluorescence protein(GFP)gene and tetracycline resistance fragment was introduced into Rhizobium sp. CZGRD1 isolated from Guangxi wild Saccharum spontaneum by a three-parent combination method. The GFP-labeled rhizobia CZGRD1 was co-cultured with Xintaitang 22(ROC22)and RB928064(B9)sugarcane cultured seedlings,and its colonization characteristics in the roots of sugarcane were examined by confocal laser microscope. The ¹⁵N isotope method was used to study the growth promotion and nitrogen fixation efficiency of CZGRD1 on B9 and ROC22 cultured seedlings and stem seedlings under potted conditions.【Result】 CZGRD1 was resistant to gentamicin and chloramphenicol,and was sensitive to ampicillin,streptomycin,spectinomycin and tetracycline. CZGRD1 invaded the roots in the root fissures,lateral root projections and root crown cells during cocultivation with B9 and ROC22 sugarcane cultured seedling. Compared with the uninoculated controls,inoculation of CZGRD1 increased plant dry weights of B9 and ROC22 sugarcane cultured seedlings by 12.11% and 4.48%,nitrogen content increased by 25.00% and 7.97%,respectively. The percentage of nitrogen fixation from air for seedlings was 12.35% and 8.30%,respectively. Inoculation of CZGRD1 increased B9 and ROC22 sugarcane stem seedlings plant dry weights by 14.31% and 10.26%,nitrogen content by 20.07% and 10.90%,respectively. The percentage of nitrogen fixation from air for seedlings was 12.77% and 10.28%,respectively.【Conclusion】Rhizobia Rhizobium sp. CZGRD1 is able to colonize in sugarcane roots,to promote the growth of B9 and ROC22 cultured seedlings and stem seedlings,and to fix nitrogen in association with sugarcane.