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We have purified and characterized two class-II restriction endonucleases from the saprophyte Mycobacterium chelonei AMB 82. Mch AI was an isoschizomer of Not I recognizing and cleaving at 5'-GC/GGCCGC. MchA I did not require Triton X-100 for activity and was fully active at concentrations over 10–200 mM KCl and MgCl 2 . MchA II was an isoschizomer of Hae II recognizing and cleaving at 5'-GG/CC. MchA II was active in 10 mM Tris-HCl pH 8.0 containing MgCl 2 at 10–200 mM, was strongly inhibited by NaCl at concentrations higher than 10 mM and was inactive at KCl concentrations lower than 250 mM.

A restriction endonuclease, MgoI, an isoschizomer of Sau3AI, was purified from Mycobacterium gordonae TRC1318. As compared to Sau3AI, the yield of MgoI was seven-eightfold higher, the enzyme was four times more stable at 37 degrees C, and in addition, had optimal activity over a much broader range of salt concentrations.