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Generation of Rat Induced Pluripotent Stem Cells (iPS) cells from rat embryonic fibroblasts

To generate germline-competent riPSCs, the researchers initially infected Wistar (WI) or Dark Agouti (DA) rat embryonic fibroblasts (REFs) from embryonic day 14.5 or 15.5 (E14.5 or E15.5) with a lentiviral vector carrying three mouse reprogramming factors (Oct3/4, Klf4, and Sox2) controlled by a tetracycline-responsive regulatory element and with Ubc-promoter - driven reverse tet transactivator (rtTA) and EGFP (Fig. 1A). Added doxycycline (Dox) to the culture medium on the day of infection (day 0), with rat LIF (rLIF) added to medium on day 1.

(A) Schema of inducible lentivirus vector. XbaI enzyme restriction site exists between Ubiquitin-C (Ubc) promoter and reverse tet transactivator (rtTA) element.

(B) Schematic time schedule of riPSCs generation.

(C)–(D) Morphology of generated riPSCs (passage P0); bright field and fluorescent field (C). ALP staining after clonal passage (D).


Links: Induced pluripotent stem cell | Stem Cells | rat

Reference

Hamanaka S, Yamaguchi T, Kobayashi T, Kato-Itoh M, Yamazaki S, Sato H, Umino A, Wakiyama Y, Arai M, Sanbo M, Hirabayashi M & Nakauchi H. (2011). Generation of germline-competent rat induced pluripotent stem cells. PLoS ONE , 6, e22008. PMID: 21789202 DOI.

Copyright

© 2011 Hamanaka et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Figure 1. Journal.pone.0022008.g001.jpg doi:10.1371/journal.pone.0022008.g001


Cite this page: Hill, M.A. (2024, June 23) Embryology Rat induced pluripotential stem cell protocol.jpg. Retrieved from https://embryology.med.unsw.edu.au/embryology/index.php/File:Rat_induced_pluripotential_stem_cell_protocol.jpg

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current14:58, 18 September 2011Thumbnail for version as of 14:58, 18 September 2011740 × 1,000 (84 KB)S8600021 (talk | contribs)==Generation of rat Induced Pluripotent Stem Cells (iPS) cells from rat embryonic fibroblasts== To generate germline-competent riPSCs, we initially infected Wistar (WI) or Dark Agouti (DA) rat embryonic fibroblasts (REFs) from embryonic day 14.5 or 15.5

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