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. 2022 Feb 14;23:54. doi: 10.1186/s13059-022-02617-x

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© The Author(s) 2022

Open AccessThis 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. 4 — The epiallelic nature of rDNA is a feature of other mouse inbred strains. A Allele-specific methylation levels at selected rDNA epivariants from the kidney of 5 inbred strains besides C57BL/6J (“BL6J”): 129S1/SvImJ (“129S”), C3H/HeJ (“C3H”), C57BL/6N (“BL6N”), CAST/EiJ (“CAST”), and A/J (“AJ”) (see Additional file 3: Fig. S13 for a comprehensive display of all identified epivariants). The top panel shows positions associated with C57BL/6J haplotypes. Notably, the methylation differences at these positions are not always directionally consistent across different strains where the variants are conserved. The bottom panel depicts strain-specific variants; note 7427 is variable in 2 strains, but with different alternative nucleotides (T in 129S1/SvImJ and G in CAST/EiJ). B Comparison of alternative allele frequencies (AAF) at the epivariant positions between rDNA (WGS) and rRNA for one representative mouse of each strain (see Additional file 3: Fig. S14). If only unmethylated units are expressed, then adjusting the AAF ratios in the WGS by ignoring methylated copies in the ratio calculations should improve the correlation between variant frequencies (i.e., unmethylated correlated with expressed). This is observed in 4/6 strains. C Total rDNA copy number from ddPCR correlated with methylation level from WGBS across the different mouse strains. Correlation coefficients displayed use all mice as data points (black) or a single average point per strain (gray). The analysis was repeated using the highly conserved 18S subunit only to exclude any possibility that the results are due to strain-specific differences in mapping efficiency (see Additional file 3: Fig. S15)