Tbsmt3 gene from the ectomychorrizal fungus Tuber borchii was identified and sequenced. The Tbsmt3 gene encodes for a protein sharing significant amino acid homology with the yeast SMT3, a ubiquitin-like protein that is post-translationally attached to several proteins involved in many cellular processes. The comparison between the Tbsmt3 genomic and cDNA sequences established that the encoding sequence is interrupted by an intron of 312 bp. Southern blot analysis revealed only one copy of Tbsmt3 gene in the T. borchii genome. Tbsmt3 is expressed in all phases of T. borchii life cycle: mycelium, ectomycorrhiza and ascoma. However, the Tbsmt3 mRNA decreased during fruit body maturation.