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. 2023 Sep 20;14:5849. doi: 10.1038/s41467-023-41506-0

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a In vitro CMG pulldown assays to examine the binding affinity of Polε, PolεΔ2N, and the related mutants (Pol2ΔDS2 and Pol2ΔDS2+4) to CMG in the presence or absence of DNA. Similar results were obtained in two independent experiments. b Schematic illustration of pol2 mutant construction. c Polε-MCM coupling is essential for cell viability. The indicated truncation mutants of Pol2 were tested for complementation of a depletion of Pol2-AID, the expression of which is under the control of Tet promoter. YPD: yeast extract-peptone-dextrose, NAA: 1-naphthalene acetic acid, Dox: doxycycline. d Flow cytometry profiles of cells collected from the α factor block-and-release assays with the indicated pol2-iAID strains under restrictive conditions. e The lysates of the indicated samples were prepared for Flag IP with anti-FLAG beads. Each sample was analyzed with antibodies against Flag, Cdc45, and Mcm2. Similar results were obtained in two independent experiments. Source data are provided as a source data file. f Flow cytometry profiles of cells collected from the hydroxyurea (HU) block-and-release assays. α factor was added after cells were released from HU arrest.