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Embryonic Stem Cells/Induced Pluripotent Stem Cells
Version of Record online: 22 OCT 2012
Copyright © 2012 AlphaMed Press
Volume 30, Issue 11, pages 2596–2601, November 2012
How to Cite
Greder, L. V., Gupta, S., Li, S., Abedin, Md. J., Sajini, A., Segal, Y., Slack, J. M.W. and Dutton, J. R. (2012), Brief Report: Analysis of Endogenous Oct4 Activation during Induced Pluripotent Stem Cell Reprogramming Using an Inducible Oct4 Lineage Label. STEM CELLS, 30: 2596–2601. doi: 10.1002/stem.1216
Author contributions: L.V.G.: conception and design, collection and/or assembly of data, data analysis and interpretation, manuscript writing, and final approval of manuscript; S.G.: conception and design, financial support, administrative support, data analysis and interpretation, and final approval of manuscript; S.L., Md.J.A., and A.S.: collection and/or assembly of data, data analysis and interpretation, and final approval of manuscript.; Y.S. and J.R.D.: conception and design, financial support, administrative support, collection and/or assembly of data, data analysis and interpretation, manuscript writing, and final approval of manuscript; J.M.W.S.: conception and design, financial support, administrative support, data analysis and interpretation, manuscript writing, and final approval of manuscript.
Disclosure of potential conflicts of interest is found at the end of this article.
First published online in STEM CELLSEXPRESS September 4, 2012.
- Issue online: 22 OCT 2012
- Version of Record online: 22 OCT 2012
- Accepted manuscript online: 4 SEP 2012 10:01AM EST
- Manuscript Accepted: 28 JUL 2012
- Manuscript Received: 30 MAR 2012
- NIH. Grant Numbers: DK078706, U01HL100407
- Minneapolis VA Health Care System
Additional Supporting Information may be found in the online version of this article.
|sc-12-0316_sm_SupplFigure1.pdf||333K||Supplementary Figure S1: Oct4-MerCreMer transgene accurately reflects endogenous Oct4 expression (A, B): Southern blot indicating correct targeting at the modified Oct4 locus. (C-F): Correlation of EGFP with endogenous Oct4 expression. Oct4-CreMerCre mT/mG fibroblasts were reprogrammed in the absence of Tamoxifen for 20 days before addition of Tamoxifen for 16 hours. Colonies were dispersed into single cells (C): before sorting (D, E): after sorting by FACS (F). (G): RT-PCR analysis shows that endogenous Oct4 gene expression is restricted to cells expressing EGFP. (H): Genomic PCR detects all 4 reprogramming viruses are integrated in both tdTomato and EGFP cell populations.|
|sc-12-0316_sm_SupplFigure2.pdf||456K||Supplementary Figure S2: Derivation and characterization of iPS cell lines from Oct4- MerCreMer; mT/mG embryonic fibroblasts reprogrammed in the absence of tamoxifen. (A): 3F-10, an iPS cell line derived after 3F OSK reprogramming, phase image. (B): 3F-10, SSEA1 staining. (C): 3F-10, tdTomato expression. (D): 3F-10, Nanog staining. (E): 7-8-8, an iPS cell line derived after 4F OSKM reprogramming, phase image. (F): 7-8-8, Oct4 staining. (G): 7-8-8, tdTomato expression. (H): 7-8-8, Nanog staining. These lines exhibit a very low background of tamoxifen independent EGFP activation common to post-translational inducible Cre systems (I): RT-PCR analysis of iPS cell lines 3F-10, 7-8-8 and 7-8-10. (J) In vitro differentiation potential during embryoid body formation. (K): Demethylation of Oct4 and Nanog promoters confirmed by bisulfite treatment and sequencing. (L-N): In vivo pluripotency determined in teratoma derived from 3F10 iPS cells, from left to right, neural tissue, muscle, and intestine-like epithelial tissue with goblet cells.|
|sc-12-0316_sm_SupplTable1.pdf||12K||Table S1: Primers used in this study|
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