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. 2022 May 5;13:2464. doi: 10.1038/s41467-022-30022-2

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© The Author(s) 2022

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. 1 — a Schematic of the mouse Dlk1-Dio3 imprinted locus showing reporter insertion. Three differentially methylated regions (DMRs) that regulate imprinted expression of the cluster are indicated (closed circles represent methylated CpGs, IG-DMR, Dlk1 sDMR and Gtl2 sDMR) and the position of maternally expressed (light grey) and paternally expressed (blue) genes are shown. Arrows depict transcriptional direction, with solid lines representing protein-coding genes and striped lines representing non-coding transcripts. In the Dlk1-FLucLacZ reporter line, firefly Luciferase (FLuc) and β–galactosidase (LacZ) were knocked into the endogenous Dlk1 locus, with T2A sites, downstream of exon 5. b Bioluminescence (BL, blue) was detected in 8-week-old (P56) male (lower panel, left) and female mice (lower panel, right) after paternal transmission of the reporter (KI^pat). Strong BL signal was evident in the thymus, central sternum and testes. Minimal signal was detected in animals after maternal reporter transmission (KI^mat, upper panel, right) or in wild-type animals (wt, upper panel, left). c Dlk1 expression analysed by QRT-PCR (upper panel) was compared in different tissues from P56 male mice that inherited the reporter paternally (KI^pat, dark grey), maternally (KI^mat, light grey), or in non-transgenic controls (wt, black). Uterus samples from age-matched female mice were also analysed. Expression levels were normalized to β-Actin, 18S and Hprt expression (bars show the geometric mean of relative expression, error bars represent the geometric standard deviation (geometric SD)). Genotype had no significant effect on Dlk1 expression (Two-way ANOVA on delta-Ct values (Tissue p < 0.0001, Genotype p = 0.86, Interaction p = 0.98); N = 4 + 4 + 4 individual mice). Allelic Dlk1 analysis in KI^mat mice (lower panel), using primers that distinguish the reporter from the wt allele, showed a strong bias for paternal allele expression (dark grey) compared to maternal allele expression (light grey). (Bars indicate the mean contribution from each allele ±SD; N = 4 + 4 individual mice). Source data are provided as a Source Data file.