Sse8387l, a useful eight base cutter for mammalian genome analysis (influence of methylation on the activity of Sse83871)

摘要

To develop restriction enzymes that are useful for genome analysis, we previously performed screening and isolated Sse8387l from Streptomyces sp. strain 8387. Sse8387l is a restriction enzyme that recognizes 5'-CCTGCA/GG-3' and cleaves DNA at the siteshown by the diagonal (Nucleic Acid Res., 18, 5637-5640). The present study evaluated the effects of methylation that is important when Sse8387l is used for genome analysis. Sse8387l lost cleavage activity after methylation of adenine or methylation ofcytosine at any site in the recognition sequence. However, the recognition sequence of Sse8387l contains no CG sequence, which is the mammalian methylation sequence. In addition, we evaluated the effects of methylation of CG at sites other than the recognition sequence. The cleavage activity of Sse8387l was maintained even when CG sequences were present immediately before or after, or near the recognition sequence, and cytosine was methylated. These results suggest that CG methylation does not affect the cleavage activity of Sse8387l. Therefore, Sse6387l seems to be very useful for mammalian genome analysis.