The aim of the current study was to evaluate the subchronic toxicity of GmDREB3 gene modified wheat in the third generation rats. SPF Wistar rats were fed with transgenic wheat diet (Gm), parental wheat diet (Jimai22) and AIN-93 rodent diet (Control), respectively, for two generations, to produce the third generation rats which were used for this study. The selected fresh weaned offspring rats (20/sex/group) were given the same diet as their parents for 13 weeks. No toxicity-related changes were observed in rats fed with Gm diet in the following respects: clinical signs, body weights, body weight gains, food consumption, food utilization rate, urinalysis, hematology, serum biochemistry and histopathology. The results from the present study demonstrated that 13 weeks consumption of Gm wheat did not cause any adverse effects in the third generation rats when compared with the corresponding Jimai22 wheat.