Background/Question/Methods In order to understand the evolutionary factors that influence ecologically important trait variation we have cloned a QTL controlling insect resistance in natural plant populations. Boechera stricta is a perennial, genetically tractable, ecological model organism, which grows in undisturbed environments. We used positional cloning to identify the genetic basis of metabolic variation in glucosinolate compounds controlling insect resistance, followed by transgenic verification of the cloned QTL. In addition, we quantified insect resistance and plant fitness using near-isogenic lines in the original environments where these alleles evolved.Results/Conclusions This QTL encodes an early enzyme in glucosinolate biosynthesis. The ancestral allele occurs in a population with low levels of herbivory, where resistance is unrelated to fitness. In contrast, the derived allele comes from a high-herbivory environment, where strong natural selection favors plants that are resistant to insect herbivores. In addition to this study of ongoing natural selection, polymorphism in this ecologically important gene will elucidate historical patterns of natural selection.