Transgenic human ES and iPS reporter cell lines for identification and selection of pluripotent stem cells in vitro

Ovchinnikov, Dmitry A., Titmarsh, Drew M., Fortuna, Patrick R. J., Hidalgo, Alejandro, Alharbi, Samah, Whitworth, Deanne J., Cooper-White, Justin J. and Wolvetang, Ernst J. (2014) Transgenic human ES and iPS reporter cell lines for identification and selection of pluripotent stem cells in vitro. Stem Cell Research, 13 2: 251-261. doi:10.1016/j.scr.2014.05.006

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Author Ovchinnikov, Dmitry A.
Titmarsh, Drew M.
Fortuna, Patrick R. J.
Hidalgo, Alejandro
Alharbi, Samah
Whitworth, Deanne J.
Cooper-White, Justin J.
Wolvetang, Ernst J.
Title Transgenic human ES and iPS reporter cell lines for identification and selection of pluripotent stem cells in vitro
Formatted title
Transgenic human ES and iPS reporter cell lines for identification and selection of pluripotent stem cells in vitro
Journal name Stem Cell Research   Check publisher's open access policy
ISSN 1873-5061
1876-7753
Publication date 2014-09-01
Sub-type Article (original research)
DOI 10.1016/j.scr.2014.05.006
Open Access Status DOI
Volume 13
Issue 2
Start page 251
End page 261
Total pages 11
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Language eng
Formatted abstract
Highlights
• Stable selectable pluripotency reporter lines in hES and transgene-free iPS cells.
• More sensitive pluripotency readout than conventional surface markers.
• Pluripotency enrichment facilitates directed differentiation protocols.
• Reveals heterogeneity in conventional embryoid body assays.

Optimization of pluripotent stem cell expansion and differentiation is facilitated by biological tools that permit non-invasive and dynamic monitoring of pluripotency, and the ability to select for an undifferentiated input cell population. Here we report on the generation and characterisation of clonal human embryonic stem (HES3, H9) and human induced pluripotent stem cell lines (UQEW01i-epifibC11) that have been stably modified with an artificial EOS(C3+) promoter driving expression of EGFP and puromycin resistance-conferring proteins. We show that EGFP expression faithfully reports on the pluripotency status of the cells in these lines and that antibiotic selection allows for an efficient elimination of differentiated cells from the cultures. We demonstrate that the extinction of the expression of the pluripotency reporter during differentiation closely correlates with the decrease in expression of conventional pluripotency markers, such as OCT4 (POU5F1), TRA-1-60 and SSEA4 when screening across conditions with various levels of pluripotency-maintaining or differentiation-inducing signals. We further illustrate the utility of these lines for real-time monitoring of pluripotency in embryoid bodies and microfluidic bioreactors.
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

 
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Created: Wed, 16 Jul 2014, 02:38:53 EST by Dr Dmitry Ovchinnikov on behalf of Aust Institute for Bioengineering & Nanotechnology