Thermal conditions are an important determinant of performance and species distributions, and the ability to cope with temperature change depends on an organism’s genotype, previous exposure to thermal environments, and the combination of these two factors. Here, we use a subset of isogenic lines from the Drosophila Genetic Reference Panel (DGRP) to investigate genotype by environment interactions in thermal limits. To test for developmental and adult acclimation, individuals from each DGRP line were kept at standard (25°C), hot (30°C), or cool (18°C) conditions during larval development or as adults, respectively, and we also tested for short-term hardening capacity by rearing flies at standard conditions and subjecting them to an acute hardening treatment immediately before testing. We then measured critical thermal minimum (CT_min) and heat knockdown time of these flies using high throughput methods developed by our group. Preliminary results suggest that the adult thermal environment and acute hardening have strong effects on thermal tolerance, while developmental conditions have a weak effect. In ongoing experiments, we are testing these conditions across lines, so that we can quantify genetic variation in thermal acclimation capacity. Results from this study will identify DGRP lines that have particularly plastic or rigid thermal limits in response to their thermal environment. In the future, this information will be used in a genome-wide association study to characterize the genetic architecture underpinning variation in thermal acclimation capacity and to identify the proximate molecular mechanisms that drive this variation.