Direct Quantitative Bisulfite Sequencing Using Tag-modified Primers and Internal Normalization

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Figure 1.
Principle of signal normalization. Sequence electropherograms (cytosine traces only) from five mixtures of methylated and unmethylated DNA (0, 5, 15, 50, and 100% methylated DNA) before (A) and after (B) signal normalization. Normalization is achieved by dividing the intensity by the area of the normalization signal.
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Figure 2.
Correlation of normalized peak areas and relative methylation of the template DNA. Normalized peak areas are shown of five single CpG sites resulting from the sequencing of seven different mixtures of methylated and unmethylated DNA (0, 5, 10, 25, 50, 75 and 100% methylated DNA). The normalized and integrated intensities are scaled from 0% to 100% in order to enable a direct comparison of the five single CpG sites.