Raw gene expression data for "Gene expression and drought response in an invasive thistle." To complement studies of phenotypic variation, we investigate gene expression during drought response that might underlie variation in drought tolerance between native and introduced populations of diffuse knapweed (Centaurea diffusa, Asteraceae), and possibly contribute to invasion success. Using species-specific microarrays and tissue sampled under drought and control conditions at three time points, we identified genes whose expression either varied constitutively or responded to drought stress differently between ranges. Further, we functionally investigate these genes. Raw expression data: In collaboration with NimbleGen, we developed a high-density customized expression 12-plex array with 60-mer probes from the transcriptomes of two C. diffusa individuals, from one native (population TR001) and one invasive (population US022) individual (Lai et al. 2012). In total, 61,024 unigenes were represented on the array with two or three probes per unigene for a total of 136,906 features. We prepared the double-stranded cDNA according to the array manufacturer’s instructions (Roche NimbleGen; NimbleGen Arrays User’s Guide v.5.1). Following Hodgins et al. (2013), we added 1 µL SUPERase (Ambion) to the first-strand synthesis reaction to prevent RNA degradation. We obtained cDNA yields with both a Nanodrop and a Qubit 2.0 Fluorometer and assessed cDNA quality using an Agilent 2100 Bioanalyzer. We labeled cDNA and performed the hybridization on a 12-bay NimbleGen Hybridization System, using the array washes recommended by NimbleGen (NimbleGen Arrays User Guide v.5.1). Using a GenePix Professional 4200A scanner (Axon Instruments), we acquired individual array images independently adjusting the PMT gain for each image as recommended.