Fig. 1: 6-base-CUT&Tag conceptualisation.
From: Sequencing DNA methylation and hydroxymethylation at co-occurring chromatin features
a Scheme depicting cytosine (C) methylation by DNA methyltransferase (DNMT) to form 5mC and progressive oxidation by (Ten-Eleven Translocation) TET enzymes to generate 5hmC, 5fC and 5caC modifications. Both 5fC and 5caC can be excised by thymine DNA glycosylase (TDG) to restore C and complete the demethylation cycle. Chemical groups specific to each cytosine state are flagged in red. b Conceptual overview illustrating how ‘standard’ whole-genome 6-base sequencing reports an average ensemble picture that does not accurately represent co-occurring epigenetic features. In contrast, enriching for a specific chromatin feature (e.g., H3K27ac mark) isolates a sub-family of co-occurring protein-DNA fragments and simultaneously reports on 5mC and 5hmC at read level. For simplicity, methylation heterogeneity for each modification is not represented in this scheme. Schematic 5mC and 5hmC percentages are shown for the CpG sites in each case. c Complete 6-base-CUT&Tag (6B-C&T) workflow. Specific steps are detailed in the main text (also see Supplementary Fig. 1). An example profile for H3K27ac, 5mC and 5hmC is shown at the bottom of the diagram and exemplifies the coupling of three layers of information originating from the same DNA fragment. Colour code for selected nucleobases is indicated throughout the diagram.
