PiggyBac transposon-mediated cellular transgenesis in mammalian forebrain by in utero electroporation

Fuyi Chen, Brady J. Maher, Joseph J. Loturco

Research output: Contribution to journalArticle

Abstract

In utero electroporation (IUE) is an effective transfection method for delivering plasmid DNA into neural progenitor cells and neurons of mammalian neocortex in vivo. Although IUE is effective at delivering multiple DNA plasmids into populations of cells, unfortunately plasmids delivered into neural progenitor cells remain largely episomal and often get inactivated or lost after cell division. This results in a form of “birthdate” labeling in which only the cell types that do not undergo a second cell division continue to express the transfected plasmids. This limits the application of IUE with standard plasmids and precludes its use in experiments where manipulating or labeling the complete cell lineage of a progenitor is desired. To circumvent this episomal loss of plasmid in IUE, we have used a binary piggyBac transposon system to induce nonviral genomic integration of transgenes. These transgenes do not appear to inactivate after cell division, and this results in stable somatic cellular transgenesis of neurons and glia. Like standard IUE, the system can be used with multiple combinations of plasmids to achieve multicolor labeling and both loss-of-function and gain-of-function manipulations. In this protocol, we describe the method for delivering a binary piggyBac transposon plasmid system by IUE.

Original languageEnglish (US)
Pages (from-to)741-749
Number of pages9
JournalCold Spring Harbor Protocols
Volume2014
Issue number7
DOIs
StatePublished - Jul 1 2014
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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