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Efficient recombinase mediated cassette exchange in hPSC to study the hepatic lineage reveals AAVS1 locus-mediated transgene inhibition
Journal Contribution - Journal Article
Tools for rapid and efficient transgenesis in “safe harbour”•loci in an isogenic context remain important to fully exploit the possibilities of human pluripotent stem cells (hPSC). We created hPSC master cell lines suitable for FLPe recombinase-mediated cassette exchange (RMCE) in the AAVS1 safe harbour locus. RMCE using positive/negative selection allows generation of transgenic lines in 15 days with 100% targeting efficiency and without random integrations. Using RMCE, we successfully incorporated a number of transgenes useful for lineage identification, assessment of cell toxicity, and gene over-expression. However, we observed unexpected and variable inhibition of expression of some of the transgenes in the AAVS1 locus, caused by DNA methylation and other unknown mechanisms, in undifferentiated hESC and during hepatic differentiation. Therefore, the AAVS1 locus cannot be considered a universally safe harbour locus, at least for the hepatic lineage, and using it for transgenesis in human cells will require thorough assessment of promoter function.
Journal: Stem Cell Reports
ISSN: 2213-6711
Issue: 2
Volume: 10
Pages: 673 - 673
Publication year:2015
- See also: Erratum: Efficient Recombinase-Mediated Cassette Exchange in hPSCs to Study the Hepatocyte Lineage Reveals AAVS1 Locus-Mediated Transgene Inhibition (Stem Cell Reports (2015) 5(5) (918–931)(S2213671115002660)(10.1016/j.stemcr.2015.09.004))
- See also: Efficient Recombinase-Mediated Cassette Exchange in hPSCs to Study the Hepatocyte Lineage Reveals AAVS1 Locus-Mediated Transgene Inhibition
- See also: Efficient Recombinase-Mediated Cassette Exchange in hPSCs to Study the Hepatocyte Lineage Reveals AAVS1 Locus-Mediated Transgene Inhibition