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. 2019 Feb 22;10:910. doi: 10.1038/s41467-019-08886-8

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© The Author(s) 2019

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 6 — Increased levels of Claspin and Timeless promote fork progression in cells expressing oncogenic Ras. a Protein levels of Claspin and Timeless in clones #4 and #5 transfected with control (si-Ctrl), Claspin (si-CLSPN), and Timeless (si-TIM) siRNAs. b DNA fiber spreading analysis of fork progression in clones #4 and #5 after depletion of Claspin and Timeless with siRNAs. Median CldU tracks length (red) were determined for three independent experiments. ****p < 0.0001, ***p < 0.001, ns nonsignificant difference. Mann–Whitney rank sum test. c Levels of Claspin and Timeless in U2OS-CycE cells transfected with GFP, Claspin, and Timeless plasmids. U2OS-CycE cells expressing Cyclin E under the control of a doxycyclin-inducible promoter were grown during 6 days in the presence of Dox to induce replication stress. Then cells are transfected with plasmids to overexpress GFP, Claspin or Timeless during 3 days before performing DNA spreading experiment. d DNA spreading analysis of experiment performed in c. DNA spreading was also performed on U2OS-CycE cells growing without Dox. Median CldU tracks length (red) were determined for three independent experiments. ****p < 0.0001, ns nonsignificant. Mann–Whitney rank sum test