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. 2023 Aug 4;21:167. doi: 10.1186/s12915-023-01669-0

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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/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 1 — Inducible depletion of SPT16 triggers reduced pluripotency in ES cells. A Schematic of auxin-inducible degron (AID) and V5-tagged SPT16 protein. NTD = N-terminal domain, DD = dimerization domain, MD = middle domain, CTD = C-terminal domain, AID = minimal auxin-inducible degron tag, V5 = 3xV5 epitope tag. B Western blot showing depletion of SPT16 after 0, 3, 6, 24, and 48 h treatments with IAA (+) or vehicle control (EtOH, −). 40 µg total protein loaded per lane. Top: V5 antibody (for tagged SPT16) and bottom: β-actin antibody. Representative blot shown from SPT16-V5-AID clone 1; additional blots can be found in Additional File 1: Fig. S1. “M” denotes a molecular weight marker lane. C Timecourse of IAA or EtOH treatment for 6, 12, 24, or 48 h to deplete SPT16 showing morphological changes following alkaline phosphatase staining. Images are representative of plate-wide morphological changes. Alkaline phosphatase staining is quantified in Additional File 1: Fig. S1E. D. Timecourse of IAA or EtOH treatment for 12, 24, or 48 h to deplete SPT16, followed by immunocytochemistry showing a progressive reduction in the expression of OCT4. Images are representative of plate-wide immunofluorescence changes. Left panels are DAPI-stained (blue), while right panels show OCT4 immunofluorescence (green). OCT4 immunofluorescence is quantified in Additional File 1: Fig. S1F