Figure 4

Sequencing analysis of dMS-HRM products from an acute myeloid leukaemia sample. PCR clones from sample 9164 were selected for sequencing as it showed the most variation, and was therefore the best example of the relationship of peak position to the degree of methylation. One representative of each peak observed from dMS-HRM is shown (a). Peaks are grouped within colours according to the number of CpG dinucleotides methylated as shown by sequencing and represented by lollipops, where open and filled circles represent unmethylated and methylated CpG sites, respectively (b). The order of clones in panel (b) reflects the order of peaks from left to right. The asterisk (*) marks PCR clone 69, as although it showed complete methylation, it also contained an unconverted cytosine. This is shown in the sequencing trace in the centre panel of (c), compared with an unmethylated sequence (above) and a methylated sequence (below). The incompletely converted cytosine is indicated in red. This incomplete conversion explains the higher Tm of this clone with respect to the other clones showing complete methylation.