Abstract The diversity of cacao (Theobroma cacao L.) genetic resources can be increased through mutation breeding. The study aimed to determine the LD20 and LD50 information of gamma-ray irradiation on cacao seeds. The research was conducted from January to December 2020 in the greenhouse and the Integrated Laboratory of The Indonesian Industrial and Beverage Crops Research Institute (IIBCRI). The cacao seeds were obtained from a prospective mother garden belonging to the IIBCRI. Seeds were applied at the Isotope and Radiation Application Center (PAIR) BATAN. Five bulk cacao clone seeds, including Sulawesi 1, Sulawesi 2, MCC 01, MCC 02, and SCA 6, were irradiated at doses of 0, 20, 40, 60, 80, and 100 Gy. The observed characteristics include the percentage of germination, root length, number of leaves, leaf length, leaf width, petiole length, plant height, and leaf shape. Each clone exhibited different LD20 and LD50 values based on the percentage of germination. The LD20 ranged from 5.83 Gy (Sulawesi 1) to 11.79 Gy (SCA 6) and LD50 ranged from 19.58 Gy (Sulawesi 1) to 25.07 Gy (SCA 6). Morphological changes in leaf shape were also observed. Increasing the dose of gammarays decreased the average values of root length, number of leaves, leaf length, leaf width, petiole length, and plant height of five cacao clones. The optimal dose of gammaray irradiation for cacao seeds was found to be 20Gy. In the future, cacao mutants will be used as parents in a plant breeding program.