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. 2026 Feb 5;54(4):gkag089. doi: 10.1093/nar/gkag089

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© The Author(s) 2026. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 2. — Tracking of DNMT1-Halo tagged molecules by live cell imaging. (A) Cell nucleus showing Halo-tagged DNMT1 molecules labeled with 0.25 nM JF646 Halo ligand for 5 min and imaged using HiLo microscopy at ∼ 97 fps over a 10 s period. Still from representative Halo-tagged DNMT1 E8 (+/+) movie (composite of five frames, ∼ 0.5 s). Dotted line denotes outline of the nucleus, scale bar = 5 µm. (B) Representative nucleus showing DNMT1-Halo-FLAG trajectories in an asynchronous E8 (+/+) cell. Left, all trajectories shown in a 10 s movie. Right, a subset of the trajectories from the last 4.5 s of the corresponding movie. (C-F) Spot-on analysis showing the dynamics of DNMT1-Halo-FLAG tagged molecules in the E8 (+/+) and F3 (+/-) cell lines, using a three-state model. (C–E) Violin plots showing the distribution of 183-236 cells across at least five replicates. (F) Pie chart showing the distribution of fast or slow diffusing and chromatin-bound DNMT1 molecules in asynchronous E8 (+/+) cells. (G) Diffusion coefficient (D) of various chromatin-modifying factors, transcription factors and DNA repair machinery proteins imaged by single molecule live cell imaging plotted against their molecular weight (represented as inverse cubic root molecular weight , units of MW = kDa). Dotted line represents the best fit (linear regression). Generally, the diffusion coefficient decreases with increased molecular weight of the protein. Source of data given in Supplementary Table 4.

Inline graphic — Tracking of DNMT1-Halo tagged molecules by live cell imaging. (A) Cell nucleus showing Halo-tagged DNMT1 molecules labeled with 0.25 nM JF646 Halo ligand for 5 min and imaged using HiLo microscopy at ∼ 97 fps over a 10 s period. Still from representative Halo-tagged DNMT1 E8 (+/+) movie (composite of five frames, ∼ 0.5 s). Dotted line denotes outline of the nucleus, scale bar = 5 µm. (B) Representative nucleus showing DNMT1-Halo-FLAG trajectories in an asynchronous E8 (+/+) cell. Left, all trajectories shown in a 10 s movie. Right, a subset of the trajectories from the last 4.5 s of the corresponding movie. (C-F) Spot-on analysis showing the dynamics of DNMT1-Halo-FLAG tagged molecules in the E8 (+/+) and F3 (+/-) cell lines, using a three-state model. (C–E) Violin plots showing the distribution of 183-236 cells across at least five replicates. (F) Pie chart showing the distribution of fast or slow diffusing and chromatin-bound DNMT1 molecules in asynchronous E8 (+/+) cells. (G) Diffusion coefficient (D) of various chromatin-modifying factors, transcription factors and DNA repair machinery proteins imaged by single molecule live cell imaging plotted against their molecular weight (represented as inverse cubic root molecular weight , units of MW = kDa). Dotted line represents the best fit (linear regression). Generally, the diffusion coefficient decreases with increased molecular weight of the protein. Source of data given in Supplementary Table 4.