Radiotherapy-related pain is a common adverse reaction with a high incidence among cancer patients undergoing radiotherapy and remarkably reduces the quality of life. However, the mechanisms of ionizing radiation (IR)-induced pain are largely unknown. In present study, mice were treated with 20 Gy X-ray to establish IR-induced pain model. X-ray evoked a prolonged mechanical, heat and cold allodynia in mice. Transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) were significantly upregulated in lumbar dorsal root ganglion (DRG). The mechanical and heat allodynia could be transiently reverted by intrathecal injection of TRPV1 antagonist capsazepine and TRPA1 antagonist HC-030031. Additionally, the phosphorylated ERK and JNK in pain neural pathway were induced by X-ray treatment. Our findings indicated activation of TRPA1 and TRPV1 is essential for the development of X-ray-induced allodynia. Furthermore, our findings suggest targeting on TRPV1 and TRPA1 may be promising prevention strategies for X-ray-induced allodynia in clinical practice.