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. 2020 Nov 27;11:6053. doi: 10.1038/s41467-020-19879-3

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© The Author(s) 2020

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 Genomic location of Firre and CrossFirre (Gm35612) on the mouse X chromosome (UCSC mm10 build 38 browser tracks). Note that Firre has multiple alternative transcripts. The locations of the CRISPR guide RNAs used to edit the locus (cut), and of the RT-PCR primer pairs to specifically detect Firre expression (F/R), strand-specific expression of CrossFirre (F1/R1), or strand-specific expression in a region of overlap between Firre and CrossFirre (F2/R2) are indicated. b RT-PCR analysis using the F/R primer pair detects Firre expression in WT, ΔFirre^Xi, and InvFirre^Xi, but not in ΔFirre^Xa. Firre expression was measured in n = 3 biologically independent samples per cell type. c Sanger sequencing analyses of a CrossFirre region (F1/R1) and of a region of overlap between Firre and CrossFirre (F2/R2) confirm heterozygosity of SNPs (BL6 on the Xi and spretus on the Xa) in each region assayed. Genomic DNA (gDNA) shows heterozygosity at the SNPs, while cDNA only shows expression from the spretus SNP (Xa). d Strand-specific analysis of CrossFirre and Firre was done using reverse transcription using either random primers, or F1, R1, F2, R2 primers, followed by PCR using F1/R1 or F2/R2 primer pairs. Firre and CrossFirre expression was measured in n = 3 biologically independent samples per cell type. Source data are provided as a Source Data file.