DNA with two closely spaced Msp I/ Hpa II sites in five methylation states and the expected MSAP signal. A model DNA molecule with two closely spaced CCGG sites is depicted in five different methylated variants. To simplify, only one DNA strand is shown, and a symmetrical methylation pattern is expected. Variants 1–3 are found in both vertebrate and plant DNA, whereas variants 4 and 5 are plant specific. The methylation state of the two CCGG sites (Site s and l) influences digestion by MspI and HpaII. The expected MSAP MH patterns of the generated short (S) and long (L) fragments (Frag.) are drawn in boxes for the five methylated variants. Note that digested S and L DNA fragments may be amplified by different selective primers, and, if so, they are not visible together on one MSAP profile.