Applying a gel-based proteomic approach, the dynamic changes in root proteins of drought treated Quercus ilex subsp. Ballota [Desf.] Samp. seedlings were followed. Water stress was applied on 20 day-old holm oak plantlets by water limitation for a period of 10 and 20 days, each followed by 10 days of recovery. Root proteins were extracted using trichloroacetate/acetone/phenol protocol and subjected to two-dimensional electrophoresis. Coomassie colloidal stained gel images were analysed and spot intensity data subjected to multivariate statistical analysis. Selected consistent spots in the three biological replicas, presenting significant changes under stress, were subjected to MALDI-TOF mass spectrometry (peptide mass fingerprinting and MS/MS). For protein identification, combined search was performed with MASCOT search engine over NCBInr Viridiplantae and Uniprot databases. Taxonomy Holm oak (Quercus ilex subsp. Ballota [Desf.] Samp.). Dentro de Q. ilex hay dos subespecies, ilex ilex e ilex Ballota.