Detection of 5-methylcytosine in DNA sequences

Col E1 DNA has methylated cytosine in the sequence 5'-CC*(A/T)GG-3' and methylated adenine in the sequence 5'-GA*TC-3' at the positions indicated by asterisks(*). When the Maxam-Gilbert DNA sequencing method is applied to this DNA, the methylated cytosine (5-methylcytosine) is found to be less reactive to hydrazine than are cytosine and thymine, so that a band corresponding to that base does not appear in the pyrimidine cleavage patterns. The existence of the methylated cytosine can be confirmed by analyzing the complementary strand or unmethylated DNA. In contrast, the methylated adenine (probably N6-methyladenine) cannot be distinguished from adenine with standard conditions for cleavage at adenine.