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. 2022 Sep 16;13:5432. doi: 10.1038/s41467-022-33013-5

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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 representation of the reprogramming protocol; briefly, a transgenic “reprogrammable” female mouse on a C57BL/6J genetic background (i4F-BL6) was crossed with a Mus musculus castaneus (CAST) male mouse to generate E13.5 F1 hybrid embryos from which mouse embryonic fibroblasts (MEFs) were obtained. MEFs were reprogrammed by induction of the polycistronic Yamanaka cassette (Oct4/Sox2/Klf4/c-Myc - OSKM) in the presence of doxycycline (DOX) for 12 days. Individual clones of mouse induced pluripotent stem cells reprogrammed in Knockout Serum Replacement medium (KSR-iPSCs) were picked on day 12 and expanded until approximately day 50. B Clustering analysis of the normalised RNAseq counts for all the biological triplicates of female MEFs, female (F KSR2 and F KSR4) and male (M KSR3 and M KSR5) iPSCs and TX 2i ESCs. C Expression analysis by RNAseq of a panel of pluripotent genes in female MEFs, female (F KSR2, F KSR4), male (M KSR3 and M KSR5) iPSCs and TX 2i ESCs. The graph shows the average Log2 Reads Per Kilobase per Million mapped reads (RPKM) expression values ± Standard Deviation (SD) from biological triplicates of each sample. Source data are provided as a Source Data file. D Table and representative H&E staining of teratomas after subcutaneous injection of 2 × 10⁶ cells into the flanks of NSG mice. iPSCs efficiently contribute to ectoderm, mesoderm, endoderm and occasionally trophectoderm. i Low magnification of a mature teratoma, scale bar represents 250 µm. ii Trophectoderm-derived trophoblast giant cells (black arrowhead), associated with large vascular spaces (blue arrowhead), characteristic of placental tissue. iii Ectodermal components corresponding to squamous epithelium (black arrowhead). iv Endodermal components corresponding to respiratory-type epithelium, including ciliated (black arrowhead), and mucin-producing goblet cells (blue arrowhead). v, vi, vii, Mesodermal components (black arrowhead) corresponding to muscle, cartilage, and bone, respectively; ii-vii scale bar represents 100 µm. Table summarises the successful generation of teratomas with tissues from the three germ layers from F KSR2, F KSR4, M KSR3 and M KSR5 iPSCs. Two teratomas per cell line were generated and analysed by H&E staining.