Irradiation is a well-known and safe technique for controlling stored product insect pests and food protection as an alternative to pesticides. This study was designed to estimate the effects of gamma radiation doses in the range of 50–500 Gy on two stages of Trogoderma granarium larvae (second and fourth instar) and adult (male and female) in wheat grain. When larvae in the second or fourth instar were exposed to 300 Gy, no emerging adults were detected. Although parental adults have demonstrated no emerging adult, T. granarium irradiated with 200 Gy. 200 Gy was required for the disinfestation of the Khapra beetle. In addition, the DNA of T. granarium larvae, male and female adults, was studied in all body cells after exposing them to 200 Gy. The image analysis results found a detailed analysis of the migration patterns of DNA as well as a homogeneity study of the samples. The study implies that the comet assay is a rapid, simple, and sensitive visual method to evaluate the genotoxic effect of gamma radiation. It also investigated the effect of gamma radiation in the Capacity of three antioxidant enzymes SOD, CAT, and GST, the indicator marker of oxidative stress MDA on two stages of T. granarium. There was a change but no significant difference in SOD and Catalase levels between treated and control samples in both stages. The 200 Gy dose level had no effect on the growth of wheat. Moreover, in the adult stage, MDA contents were highly significant between the control and treated sample. Therefore, we estimated that a phytosanitary irradiation dose of 200 Gy is appropriate for T. granarium.