Directed Evolution of Improved Zinc Finger Methyltransferases
Percent methylation observed at individual CpG sites at and adjacent to the (A) target site and (B) non-target site. Percentages at each CpG site were determined by bisulfite sequencing of n number of clones (n indicated at right). CpG sites are numbered sequentially from 1–48 or 1–60 based on their order in the sequencing read and thus, the figure does not indicate the distance between sites. The asterisks indicate two CpG sites at which the methylation status of one clone was judged to be indeterminate due to poor sequencing quality in this region. Black, ‘WT’ heterodimeric enzyme (KFNSE); orange, PFCSY variant; blue, CFESY variant. The target and non-target CpG sites (i.e. the two sites assessed by restriction enzyme digests) are indicated by the arrows. The length of DNA subjected to bisulfite sequencing was 548 bp for the target site and 631 bp for the non-target site.