Type II restriction endonucleases were purified by heparin-sepharose followed by ion chromatography from Treponema strains. The results indicate that in addition to frequently cutting GATC-specific restriction enzymes, the tested strains also possess rarely cutting endonucleases. The purified restriction endonucleases represent four different sequence specificities, comprising isoschizomers of DrdI, AflII, Tth111I and NdeI. The data presented show that three rumen Treponema strains possess altogether seven type II restriction-modification systems. Thus, individual Treponema strains may be considered an interesting source of multiple type II restriction enzymes.