Chen D F, Liu Q A, Chen X W, Zhao X L, Chen Y W
Institute for Molecular Biology, Nankai University, Tianjin, China.
Nucleic Acids Res. 1991 Oct 25;19(20):5703-5. doi: 10.1093/nar/19.20.5703.
The recombinant plasmid pGEM4Z-ras DNA which was methylated on dam and dcm sites outside the PvuII recognition sequence was digested with restriction endonuclease PvuII, and one of the three PvuII sites was about 16-fold less efficient to cleave than either of the other two. On the contrary, the three PvuII sites were cleaved at about the same rate on the unmethylated DNA molecule. The results show that the cleavage inhibition of the methylated DNA on the certain PvuII site was caused by methylation outside the PvuII recognition sequence. Maybe a adjacent methylated dam site *A was responsible for the less efficient cleavage. This observation suggests that methylation outside the recognition sequence may be considered a new factor in the kinetic experiment of restriction endonuclease.
在PvuII识别序列之外的dam和dcm位点甲基化的重组质粒pGEM4Z-ras DNA，用限制性内切酶PvuII进行消化，三个PvuII位点中的一个切割效率比另外两个位点中的任何一个低约16倍。相反，在未甲基化的DNA分子上，三个PvuII位点以大致相同的速率被切割。结果表明，特定PvuII位点上甲基化DNA的切割抑制是由PvuII识别序列之外的甲基化引起的。可能相邻的甲基化dam位点*A是切割效率较低的原因。这一观察结果表明，识别序列之外的甲基化可能被视为限制性内切酶动力学实验中的一个新因素。
