In order to develop a new resistance identification method of sugarcanes to smut disease,caused by the fungus Ustiliago scitaminea Sydow, and provide technical support to breeding of smut resistant sugarcanes,single teliospore of Ustiliago scitaminea Sydow was isolated and cultured, mating type of sporidia was also identified,by using a 1∶1 mixture (plus and minus) mating types as inoculum.Injection inoculation was carried out at the growing point or young stem below it in younger sugarcane seedlings,and the concentration of mating type (plus and minus) and injection volume were optimized in this study. The results showed that comparatively the most suitable mating type concentration was 5×105 sporidia/mL,the relatively appropriate injection volume was 90 μL per plant in injection inoculation method. In comparison with common dipping inoculation,injection inoculation has a shorter latent period,high incidence,and it is possible to sufficiently evaluate resistance level of host and the type of physiological race. On this basis,twenty-two sugarcane varieties (materials) were inoculated by using the newly established injection inoculation and common dipping inoculation,the results showed that there were one susceptible variety,twenty-one high susceptible varieties by using injection inoculation method;however,one resistant variety,two moderately resistant varieties,eight susceptible varieties and eleven high susceptible varieties by using common dipping inoculation method.