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The flooded environment brings about injuries to soybeans that vary depending on the adaptation ability of the genotype. Oxygen deprivation promotes the induction of the expression of genes related to glycolysis and fermentation pathways to maintain energy metabolism and, in addition to reducing-power consuming processes, act in the formation of adaptive structures and the maintenance of the redox status of the plant. The aim of this work was to evaluate the relative expression of genes related to soil flooding response in two contrasting soybean cultivars. Soybean plants of the sensitive (BRS 154) and tolerant (I27) cultivars at the V1 development stage were submitted to the flooding and control conditions (without flooding) for 0, 24, 48, and 96 hours. The relative expression of genes associated with flooding, including enolase (ENO), alcohol dehydrogenase 1 (ADH1), alanine aminotransferase 2 (ALAT2), hemoglobin 1 (GLB1), LOB41 domain-containing protein (LBD41), xyloglucan endotransglycosylase (XETP) and ascorbate peroxidase (APX2), was evaluated by means of RT-qPCR. The relative expression, in general, increased with flooding, especially in the root tissue. Cultivar I27 responded positively as observed by the expression of the maintenance genes of energy metabolism, structural changes and detoxification, suggesting the presence of three tolerance mechanisms in the flooding response.