Messenger RNA electroporation is highly efficient in mouse embryonic stem cells: successful FLPe- and Cre-mediated recombination

P Ponsaerts, J P Brown, D Van den Plas, L Van den Eeden, D R Van Bockstaele, P G Jorens, V F I Van Tendeloo, J Merregaert, P B Singh, Z N Berneman

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16 Citations (Scopus)

Abstract

Development of efficient short-term gene transfer technologies for embryonic stem (ES) cells is urgently needed for various existing and new ES cell-based research strategies. In this study, we present a highly efficient, nonviral non-DNA technology for genetic loading of mouse ES cells based on electroporation of defined mRNA. Here, we show that mouse ES cells can be efficiently loaded with mRNA encoding a green fluorescent reporter protein, resulting in a level of at least 90% of transgene expression without loss of cell viability and phenotype. To show that transgenes, introduced by mRNA electroporation, exert a specific cellular function in transfected cells, we electroporated stably transfected ES cell lines with mRNA encoding FLPe or Cre recombinase proteins in order to excise an FRT- or LoxP-flanked reporter gene. The results, as determined by the disappearance and/or appearance of a fluorescent reporter gene expression, show that FLPe and Cre recombinase proteins, introduced by mRNA electroporation, efficiently exert their function without influence on further culture of undifferentiated ES cell populations and their ability to differentiate towards a specific lineage.

Original languageEnglish
Pages (from-to)1606-10
Number of pages5
JournalGene Therapy
Volume11
Issue number21
DOIs
Publication statusPublished - Nov 2004

Keywords

  • Animals
  • Cells, Cultured
  • Electroporation
  • Flow Cytometry
  • Gene Expression
  • Green Fluorescent Proteins
  • Integrases
  • Luciferases, Firefly
  • Mice
  • RNA, Messenger
  • Recombination, Genetic
  • Stem Cells
  • Transgenes
  • Journal Article
  • Research Support, Non-U.S. Gov't

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  • Cite this

    Ponsaerts, P., Brown, J. P., Van den Plas, D., Van den Eeden, L., Van Bockstaele, D. R., Jorens, P. G., Van Tendeloo, V. F. I., Merregaert, J., Singh, P. B., & Berneman, Z. N. (2004). Messenger RNA electroporation is highly efficient in mouse embryonic stem cells: successful FLPe- and Cre-mediated recombination. Gene Therapy, 11(21), 1606-10. https://doi.org/10.1038/sj.gt.3302342