Mutation breeding has a long track record in the development of crop cultivars with improved tolerance to abiotic stresses such as heat, salinity and drought. 'Oryza sativa' L. cv IR64 is a very popular high yielding rice, but susceptible to major abiotic stresses, such as low and high temperatures, salinity and drought. We subjected IR64 to gamma irradiation and generated a mutant population (M3) with ~2,000 families. These were screened at the seedling stage for tolerance to high-temperature stress using hydroponics and controlled-environment chambers, resulting in the identification of three mutant lines showing a robust seedling phenotype. Under heat stress, higher CO2 assimilation (10-30%), higher spikelet fertility (40-45%) and higher antioxidant activity (15-20% catalase activity) confirmed superiority of the selected mutant lines over wild type plants at seedling and flowering stages. Upon exposure to salinity and drought stress, the three selected lines also exhibited better tolerance than wild type in terms of increased CO2 assimilation, stomatal conductance, transpiration and chlorophyll fluorescence. Transcript and protein abundance analyses confirmed higher constitutive levels of heat shock proteins and antioxidant enzymes in the mutant lines relative to wild type. Tolerance to multiple abiotic stresses was reflected in higher (25-30%) grain yield than wild type. It is anticipated that the mutant lines identified will be useful for developing new improved cultivars for dry and saline areas and may be exploited to dissect the molecular basis of multiple stress tolerance in crop plants.