[Aim] The resistance against Ostrinia furnacalis, Mythimna separata and Hrelicoverpa armigera and the tolerance to the target herbicide (glyphosate)were evaluated in the transgenic maize Shuangkang 12-5-21 (hybrid of transgenic cry1Ab/cry2Aj and G10ev-epsps maize Shuangkang 12-5).[Method] The larvae of O. furnacalis were put onto field-grown maize plants at 6 leaves and silking stage, M. separata were inoculated at 6 leaves stage and H. armigera at silking stage. Under natural conditions, the resistance of Shuangkang 12-5-21 to Lepidoptera pests was investigated at harvest time. The 1, 2, 4 and 6 times recommended medium dose of glyphosate were sprayed at 5 leaves stage of maize. The damage of maize were investigated at 1, 2 and 4 weeks after spraying glyphosate.[Result] At 6 leaves and silking stage, the average damage level of Shuangkang 12-5-21 caused by O. furnacalis was 1.12 and 1.07 respectively, the level caused by M. separata at 6 leaves stage and H. armigera at silking stage was 1.32 and 0.02 respectively. Shuangkang 12-5-21 showed high resistance to O. furnacalis, M. separata and H. armigera. At harvest time the number of larval tunnels, live insects and the damage rate were significantly lower than those of the control maize, with a control effect against Lepidoptera pests was >90%. Shuangkang 12-5-21 had no damage symptoms treated with recommended medium dose of glyphosate. When treated with 2 and 4 times recommended medium dose of glyphosate, Shuangkang 12-5-21 showed slight damage symptoms 1 week later while no damage symptom 2 weeks later; treated with 6 times medium dose of glyphosate, the damage rate of herbicide of Shuangkang 12-5-21 decreased from 36.00% (1 week later) to 8.00% (2 weeks later) and the new leaves grew normally. After 4 weeks, the growth of Shuangkang 12-5-21 completely returned to normal, and the plant height was not affected.[Conclusion] Transgenic maize Shuangkang 12-5-21 showed high resistance to O. furnacalis, M. separata and H. armigera and Shuangkang 12-5-21 could tolerate 4 times recommended medium dose of glyphosate.