Abstract We examined the consistency of the native diploid Nicotiana attenuata (Na)'s herbivore-induced transcriptional changes in the two allotetraploid natives, Nicotiana clevelandii (Nc) and Nicotiana quadrivalvis (Nq), which are thought to be derived from hybridizations with an ancestral Na. An analysis of nuclear-encoded chloroplast-expressed Gln synthetase gene (ncpGS) sequences found strong similarity between Nc and Na and between N. trigonophylla and the two allopolyploids. All species were elicited with methyl jasmonate (MeJA), or were wounded and treated with either water, Manduca sexta oral secretions and regurgitant (R), or the two most abundant fatty acid amino acid conjugates (F) in R to simulate herbivory. The induced transcriptional responses in all three species were compared with a cDNA microarray enriched in Na genes. Na had the fastest transcriptional responses followed by Nc and then Nq. Na's R- and F-elicited responses were more similar to those from Nq, while the MeJA- or wound-elicited responses were more consistent in Nc. Treatment of wounds with the full cocktail of elicitors found in R elicits more complex responses than does treatment with F. The species differ in their elicited JA responses, and these differences are mirrored in the expression of oxylipin genes (LOX, HPL, AOS, and α-DOX) and downstream JA-elicited genes (TD). Elicitation decreases the expression of growth-related genes in all three species. We propose that this is a valuable system to examine the modification of complex, polygenic, adaptive responses during allopolyploid speciation.