Sagawa H, Ohshima A, Kato I
Genetic Engineering Section II, Biotechnology Research Laboratories, Takara Shuzo Co. Ltd, Shiga, Japan.
Nucleic Acids Res. 1995 Jul 11;23(13):2367-70. doi: 10.1093/nar/23.13.2367.
To develop restriction enzymes that are useful for genome analysis, we previously performed screening and isolated Sse8387I from Streptomyces sp. strain 8387. Sse8387I is a restriction enzyme that recognizes 5'-CCTGCA/GG-3' and cleaves DNA at the site shown by the diagonal (Nucleic Acid Res., 18, 5637-5640). The present study evaluated the effects of methylation that is important when Sse8387I is used for genome analysis. Sse8387I lost cleavage activity after methylation of adenine or methylation of cytosine at any site in the recognition sequence. However, the recognition sequence of Sse8387I 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 Sse8387I 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 Sse8387I. Therefore, Sse8387I seems to be very useful for mammalian genome analysis.
为了开发对基因组分析有用的限制酶，我们之前进行了筛选，并从链霉菌属菌株8387中分离出了Sse8387I。Sse8387I是一种识别5'-CCTGCA/GG-3'并在对角所示位点切割DNA的限制酶（《核酸研究》，18，5637 - 5640）。本研究评估了甲基化的影响，甲基化在Sse8387I用于基因组分析时很重要。在识别序列中任何位点的腺嘌呤甲基化或胞嘧啶甲基化后，Sse8387I失去切割活性。然而，Sse8387I的识别序列不包含CG序列，而CG序列是哺乳动物的甲基化序列。此外，我们评估了识别序列以外位点的CG甲基化的影响。即使在识别序列之前、之后或附近紧邻存在CG序列且胞嘧啶被甲基化时，Sse8387I的切割活性仍得以维持。这些结果表明，CG甲基化不影响Sse8387I的切割活性。因此，Sse8387I似乎对哺乳动物基因组分析非常有用。
