Figure 3. Representative sequencing results of 75% methylated control sample using deoxyinosine- and degenerate base primers.

Sequencing results of representative libraries comparing the positional effects of deoxyinosine- and degenerate base modifications in modified primers. (a) Comparison of methylation profile of a representative amplicons, where the modification is located near the 5′ terminus of the forward primer (left panel, ‘5′CpG’), modification is located near the centre of the forward primer (middle panel, ‘Ctr CpG’), and CpG free control (right, ‘CpG-free Control). Comparison of the three sets of amplicons showed a minimal level of difference in methylation between the tree sets of amplicons. (b) Although the portion of amplicons generated from primers with the modification at either the 5′ end or centre of the forward primer appeared to be quite different when visualised, statistical analysis showed no significant difference between the two and the CpG-free control-generated amplicons. (c) Modified bases corresponding to the box and whiskers plot in (b), with corresponding boxplot shown in [ ].