Sudden death syndrome (SDS) of soybean can be caused by any of four distinct Fusarium species, with F. virguliforme and F. tucumaniae being the main casual agents in North and South America, respectively. Although the fungal tissue is largely confined to the roots, the fungus releases a toxin that is translocated to leaf tissues, in which it causes interveinal chlorosis and necrosis leading to scorching symptoms and possible defoliation. In this study, we report on an Affymetrix analysis measuring transcript abundances in resistant (PI 567.374) and susceptible (Essex) roots upon infection by F. virguliforme, 5 and 7 days postinoculation. Many of the genes with increased expression were common between resistant and susceptible plants (including genes related to programmed cell death, the phenylpropanoid pathway, defense, signal transduction, and transcription factors), but some genotype-specific expression was noted. Changes in small (sm)RNA levels between inoculated and mock-treated samples were also studied and implicate a role for these molecules in this interaction. In total, 2,467 genes were significantly changing in the experiment, with 1,694 changing in response to the pathogen; 93 smRNA and 42 microRNA that have putative soybean gene targets were identified from infected tissue. Comparing genotypes, 247 genes were uniquely modulating in the resistant host, whereas 378 genes were uniquely modulating in the susceptible host. Comparing locations of differentially expressed genes to known resistant quantitative trait loci as well as identifying smRNA that increased while their putative targets decreased (or vice versa) allowed for the narrowing of candidate SDS defense-associated genes.