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. 2017 Jul 28;7:6838. doi: 10.1038/s41598-017-07144-5

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© The Author(s) 2017

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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Differentiation of miPS cells into CSCs. (a) Schematic flow chart of CSC generation. Detailed description is in the text. Representative images of miPS cells, and CSCcmT47D and CSCcmBT549 cells. Images of unstained cells and GFP-positive cells from identical fields of view are shown side by side. Scale bars represent 200 μm. (b) RT-qPCR analysis of the CSC markers CD133, Epcam, Oct3/4, and Sox2 in CM-T47D and CM-BT549 cells after 4 weeks in CM. Gene expression levels were normalized to those of GAPDH, and relative gene expression levels in CM-T47D or CM-BT549 cells were compared with those in untreated miPS cells. (c) RT-qPCR analysis of the Klf4 and c-Myc transgenes in CM-T47D and CM-BT549 cells after four weeks in CM, and in primary CSCcmBT549 and CSCcmT47D cells generated from subcutaneous tumors. An image of an agarose gel with the RT-qPCR products demonstrates the absence of Klf4 and c-Myc expression. The data were analyzed using an unpaired two-tailed Student’s t-test and are presented as the mean ± standard deviation **P < 0.01, *P < 0.05. (d and e) T47DCSCcm (d) and BT549CSCcm (e) primary subcutaneous tumors stained with hematoxylin-eosin. Scale bars represent 300 μm.