Conservation of telomeric DNA repeat sequences has been found across evolutionarily diverse eukaryotes. Here we report on a marked telomeric sequence diversity within the budding yeast genus Saccharomyces. Cloning and sequencing of telomeric repeat units from S. castellii, S. dairensis, S. exiguus and S. kluyveri showed a length variation between 8 and 26 bp, as well as a distinct variation in the degree of homogeneity, among the species. In S. castellii and S. dairensis, TCTGGGTG constituted a majority of the telomeric repeat units. However, the character of the variant repeats differed: in S. castellii the major class of variant repeats contained additional TG dinucleotides per repeat unit, [TCTGGGTG(TG)1-3], whereas in S. dairensis the major variant repeat is the shorter, uniform sequence TCTGGG. This result suggests mechanistic differences in the action of the telomerases of these closely related yeasts. Despite their length and homogeneity differences, all the Saccharomyces telomeric sequences show a conserved core which is also shared by the Candida glabrata telomeric sequence. This evolutionary similarity may be partly explained by the preservation of a binding site for the RAP1 protein.