Sugarcane has achieved increasing importance around the world for besides being used in sugar production, shows great potential as a bioenergy crop for the production of ethanol, an alternative, environmentally clean and renewable fuel as well as for the co-generation of electricity through the burning of bagasse. However, the crop is subject to environmental stresses and water availability is the main factor that influences plant growth, development and productivity. Thus, in order to identify the genes involved in the response of sugarcane genotype to drought, a comparative gene expression profile analysis was carried out among two drought-tolerant cultivars, SP83-5073 and RB867515, and one drought-sensitive cultivar, SP86-155. Drought was started by stopping watering and samples were collect 1, 3, 5, and 10 days after. The AFLP technique was used to identify genes differentially expressed. Using 14 combinations of EcoRI/MseI selective primers, 1,316 transcript derived fragments (TDFs) expressed only in drought-tolerant cultivars under hydric stress were detected, with 630 out of them being identified in cultivar SP83-5073 and 686 in cultivar RB867515. The analysis of the gene expression profile associated with the sugarcane genotypes revealed transcript fragments similar to the genes involved in the responses of plants to stresses, such as drought. The proteins encoded by the transcripts act in hormonal signaling, in the regulation of gene transcription, in senescence, in the transposition of DNA, amino acid transport, in respiration, and carbohydrate metabolism. However, nine TDFs seen to be unknown and may have important functions in drought tolerance of sugarcane.