Culture parameters for stable expansion, genetic modification and germline transmission of rat pluripotent stem cells

Title: Culture parameters for stable expansion, genetic modification and germline transmission of rat pluripotent stem cells
Authors: Blair, K
Leitch, HG
Mansfield, W
Dumeau, C-E
Humphreys, P
Smith, AG
Item Type: Journal Article
Abstract: The ability of cultured pluripotent cells to contribute to the germline of chimaeric animals is essential to their utility for genetic manipulation. In the three years since rat embryonic stem (ES) cells were first reported the anticipated proliferation of genetically modified rat models from this new resource has not been realised. Culture instability, karyotypic anomalies, and strain variation are postulated to contribute to poor germline colonisation capacity. The resolution of these issues is essential to bring pluripotent cell‐based genetic manipulation technology in the rat to the level of efficiency achieved in the mouse. Recent reports have described various alternative methods to maintain rat ES cells that include provision of additional small molecules and selective passaging methods. In contrast, we report that euploid, germline competent rat ES and embryonic germ (EG) cell lines can be maintained by simple adherent culture methods in defined medium supplemented with the original two inhibitors (2i) of the mitogen‐activated protein kinase (ERK1/2) cascade and of glycogen synthase kinase 3, in combination with the cytokine leukaemia inhibitory factor (LIF). We demonstrate genetic modification, clonal expansion and transmission through the germline of rat ES and EG cell lines. We also describe a marked preference for full‐term chimaera contribution when SD strain blastocysts are used as recipients for either DA or SD pluripotent stem cells.
Issue Date: 15-Jan-2012
Date of Acceptance: 1-Nov-2011
URI: http://hdl.handle.net/10044/1/62065
DOI: https://dx.doi.org/10.1242/bio.2011029
ISSN: 2046-6390
Publisher: COMPANY OF BIOLOGISTS LTD
Start Page: 58
End Page: 65
Journal / Book Title: BIOLOGY OPEN
Volume: 1
Issue: 1
Copyright Statement: © 2011 The Authors. Published by The Company of Biologists Ltd This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Share Alike License (http://creativecommons.org/licenses/by-nc-sa/3.0/).
Keywords: Science & Technology
Life Sciences & Biomedicine
Biology
Life Sciences & Biomedicine - Other Topics
rat
embryonic stem cell
embryonic germ cell
2i
chimaera
germline transmission
2i
chimaera
embryonic germ cell
embryonic stem cell
germline transmission
rat
Science & Technology
Life Sciences & Biomedicine
Biology
Life Sciences & Biomedicine - Other Topics
rat
embryonic stem cell
embryonic germ cell
2i
chimaera
germline transmission
Publication Status: Published
Online Publication Date: 2011-11-02
Appears in Collections:Clinical Sciences
Imaging Sciences
Faculty of Medicine



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