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. 2023 Mar 2;55(3):484–495. doi: 10.1038/s41588-023-01324-y

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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 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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Extended Data Fig. 1 — (a) Schematic of MERVL indicating the positions of ASOs. Intact MERVL encodes the retroviral proteins; Gag (Group-specific antigens, comprised of MA, matrix; CA, capsid proteins) and Pol (Polymerase, comprised of RT, reverse transcriptase; INT, integrase; dUTPase, dUTP phosphatase). Below whisker plot showing alignment quality in each interspersed genomic MERVL copy (adapted from Dfam: https://dfam.org/family/DF0003918/seed). (b) Pie chart indicates the populations of full length (≥ 5001 bp) and truncated (1–5000 bp) MERVL copies across the mouse genome. (c) Chromosome maps showing the distribution of MERVL copies that are targeted by ASOs throughout the mouse genome. Each colored rectangle (blue for ASO-#1, yellow for ASO-#2, and red for ASO-#3) indicates targeted MERVL copy. (d) Histogram showing the distributions of ASO-targeted (magenta) and untargeted (green) MERVL copies. The abundance of each length of MERVL element was tallied with a given count. The proportion of full-length MERVL (>5 kb) was highlighted in red. (e) Schematic for ASO-mediated KD against MERVL in ESC harboring a doxycycline (Dox)-inducible Dux transgene (termed ESC^DUX). TRE, tetracycline responsive element; rtTA, reverse tetracycline responsive transcriptional activator. (f) Expression levels of MERVL mRNA measured by qRT-PCR in ESC^DUXs nucleofected with each individual MERVL-targeting ASO and scrambled ASO. Relative expression is quantified with ΔΔCt method and normalized to Gapdh expression. Bars show means with s.e.m. Dots represent biological replicates (n = 3 samples). (g) Expression level of MERVL mRNA measured by qRT-PCR in scrambled control and ASOs (Mixed)-mediated MERVL-KD ESC^DUXs in the presence or absence of Dox. Relative expression is quantified with ΔΔCt method and normalized to Gapdh expression. Bars show means with s.e.m. Dots represent biological replicates (n = 4 samples). (h) Expressions of MERVL retroviral protein (Gag), measured by western blotting in scrambled control and ASOs (Mixed)-mediated MERVL-KD ESC^DUXs in the presence or absence of Dox. β-Tubulin was used as a loading control. Representative blot image is from 3 independent experiments. Data for panels in b-d and f-h are available as source data.

Source data