In order to elucidate the biochemical differences of the sheath blight pathogens among rice (Oryza sativa L.),corn (Zea mays L.) and wheat (Triticum aestivum L.),the total soluble proteins and esterase isozymes from 18 strains of Rhizoctonia spp. collected from these three crops were analyzed using vertical slab polyacrylamide gel electrophoresis. The results showed that the patterns of total soluble proteins and esterase isozymes of Rhizoctonia strains from three crops were significantly different,of which total soluble protein were more diverse. The band numbers of total soluble proteins from rice and corn strains were 14 to 19,and those from wheat strains were 7 to 9. The band numbers of esterase isozymes from rice and corn strains were 5 to 7,and those from wheat strains were only 3，indicating obvious differences among them. The results of cluster analysis based on NTSYS software showed that the profiles of total soluble proteins and esterase isozymes of Rhizoctonia spp. from rice,corn and wheat had a rich diversity. These findings will lay a foundation for the biochemical identification of the crop sheath blight pathogens.