To study the evolution of the polymeric β-fructosidase (invertase) genes (SUC) of yeasts Saccharomyces, new SUC gene of S. cariocanus was cloned and sequenced and the nucleotide and amino acid sequences were compared for all known β-fructosidases of Saccharomyces species. The proteins showed 90–97% homology. The most divergent was S. bayanus β-fructosidase. The results testified again to high conservation of yeast β-fructosidases. Transitions C-T prevail in the total spectrum of nucleotide substitutions observed in the coding regions of the SUC genes; most of these transitions are in the third codon position and cause no changes in the amino acid sequences of the encoded proteins. The six Saccharomyces species each carry one (probably, non-telomeric) β-fructosidase gene. SUC is on chromosome IX in S. cerevisiae, S. bayanus, S. kudriavzevii, S. mikatae, and S. paradoxus and in a translocation region on chromosome XV in S. cariocanus.