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Please use this identifier to cite or link to this item: http://hdl.handle.net/1942/20238

Title: Efficient recombinase-mediated cassette exchange in hPSCs to study the hepatocyte lineage reveals AAVS1 locus-mediated transgene inhibition
Authors: Ordovás, Laura
Boon, Ruben
Pistoni, Mariaelena
Chen, Yemiao
Wolfs, Esther
Guo, Wenting
Sambathkumar, Rangarajan
Bobis-Wozowicz, Sylwia
Helsen, Nicky
Vanhove, Jolien
Berckmans, Pieter
Cai, Qing
Vanuytsel, Kim
Eggermont, Kristel
Vanslembrouck, Veerle
Schmidt, Béla Z.
Raitano, Susanna
Van den Bosch, Ludo
Nahmias, Yaakov
Cathomen, Toni
Struys, Tom
Verfaillie, Catherine M.
Issue Date: 2015
Citation: Stem Cell Reports, 5 (5), p. 918-931
Abstract: Tools for rapid and efficient transgenesis in “safe harbor” loci in an isogenic context remain important to exploit the possibilities of human pluripotent stem cells (hPSCs). We created hPSC master cell lines suitable for FLPe recombinase-mediated cassette exchange (RMCE) in the AAVS1 locus that allow generation of transgenic lines within 15 days with 100% efficiency and without random integrations. Using RMCE, we successfully incorporated several transgenes useful for lineage identification, cell toxicity studies, and gene overexpression to study the hepatocyte lineage. However, we observed unexpected and variable transgene expression inhibition in vitro, due to DNA methylation and other unknown mechanisms, both in undifferentiated hESC and differentiating hepatocytes. Therefore, the AAVS1 locus cannot be considered a universally safe harbor locus for reliable transgene expression in vitro, and using it for transgenesis in hPSC will require careful assessment of the function of individual transgenes.
Notes: Correspondence: laura.ordovas@med.kuleuven.be or lordovas@unizar.es (L.O.), catherine.verfaillie@med.kuleuven.be (C.M.V.)
URI: http://hdl.handle.net/1942/20238
DOI: 10.1016/j.stemcr.2015.09.004
ISI #: 000364991000021
ISSN: 2213-6711
Category: A1
Type: Journal Contribution
Validation: ecoom, 2016
Appears in Collections: Research publications

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