The objective of our study was to assess the radioprotective effect of flavonoids extracted from Rosa roxburghii Tratt (FRT) and investigate the role of Bcl-2(Ca 2+ )/Caspase-3/PARP-1 pathway in radiation-induced apoptosis. Cells and mice were exposed to 60 Co γ-rays at a dose of 6 Gy. The radiation treatment induced significant effects on tissue pathological changes, apoptosis, Ca 2+ , ROS, DNA damage, and expression levels of Bcl-2, Caspase-3 (C-Caspase-3), and PARP-1. The results showed that FRT acted as an antioxidant, reduced DNA damage, corrected the pathological changes of the tissue induced by radiation, promoted the formation of spleen nodules, resisted sperm aberration, and protected the thymus. FRT significantly reduced cell apoptosis compared with the irradiation group. The expression of Ca 2+ and C-Caspase-3 was decreased after FRT treatment compared with the radiation-treated group. At the same time, expression of prototype PARP-1 and Bcl-2 increased, leading to a decrease in the percentage of apoptosis cells in FRT treatment groups. We conclude that FRT acts as a radioprotector. Apoptosis signals were activated via the Bcl-2(Ca 2+ )/Caspase-3/PARP-1 pathway in irradiated cells and FRT inhibited this pathway of apoptosis by down-regulation of C-Caspase-3 and Ca 2+ and up-regulation of prototype PARP-1 and Bcl-2.