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The sequence specificity of an extensively purified DNA methylase preparation from Krebs II mouse ascites cells has been examined. The enzyme appears to be highly sequence dependent. Moreover the sequence distribution of cytosine residues that are methylated, bears a very close resemblance to the sequence distribution of 5'-methyl cytosine found in vivo in a wide range of vertebrate cells and is consistent with methylation of cytosines in the sequence R-Yn-C-R.