Powdery mildew is an economically important disease in a number of crop legumes; however, little is known about resistance to the disease in these species. To gain a better understanding of the genetics of resistance and plant responses to powdery mildew in legumes, we developed a pathosystem with Medicago truncatula and Erysiphe pisi. Screening accessions of M. truncatula identified genotypes that are highly susceptible, moderately resistant and highly resistant to the fungus. In the highly resistant genotype, fungal growth was arrested after appressorium development with no colony formation, while in the moderately resistant genotype a small number of colonies formed. Both resistant and moderately resistant genotypes produced hydrogen peroxide and fluorescent compounds at pathogen penetration sites, consistent with a hypersensitive response (HR), although the response was delayed in the moderately resistant genotype. Very little hydrogen peroxide or fluorescence was detected in the susceptible accession. Microarray analysis of E. pisi-induced early transcriptional changes detected 55 genes associated with the basal defence response that were similarly regulated in all three genotypes. These included pathogenesis-related genes and other genes involved in defence, signal transduction, senescence, cell wall metabolism and abiotic stress. Genes associated with the HR response included flavonoid pathway genes, and others involved in transport, transcription regulation and signal transduction. A total of 34 potentially novel unknown genes, including two legume-specific genes, were identified in both the basal response and the HR categories. Potential binding sites for two defence-related transcription regulators, Myb and Whirly, were identified in promoter regions of induced genes, and four novel motifs were found in promoter regions of genes repressed in the resistant interaction.