Introducing gene deletions by mouse zygote electroporation of Cas12a/Cpf1

Charles Etienne Dumeau*, Asun Monfort, Lucas Kissling, Daan C. Swarts, Martin Jinek, Anton Wutz

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

1 Citation (Scopus)

Abstract

CRISPR-associated (Cas) nucleases are established tools for engineering of animal genomes. These programmable RNA-guided nucleases have been introduced into zygotes using expression vectors, mRNA, or directly as ribonucleoprotein (RNP) complexes by different delivery methods. Whereas microinjection techniques are well established, more recently developed electroporation methods simplify RNP delivery but can provide less consistent efficiency. Previously, we have designed Cas12a-crRNA pairs to introduce large genomic deletions in the Ubn1, Ubn2, and Rbm12 genes in mouse embryonic stem cells (ESC). Here, we have optimized the conditions for electroporation of the same Cas12a RNP pairs into mouse zygotes. Using our protocol, large genomic deletions can be generated efficiently by electroporation of zygotes with or without an intact zona pellucida. Electroporation of as few as ten zygotes is sufficient to obtain a gene deletion in mice suggesting potential applicability of this method for species with limited availability of zygotes.

Original languageEnglish
Pages (from-to)525-535
JournalTransgenic Research
Volume28
Issue number5-6
Early online date3 Sep 2019
DOIs
Publication statusPublished - Dec 2019

Keywords

  • Cas12a
  • Cpf1
  • CRISPR-Cas
  • Electroporation
  • Gene deletion
  • Mouse embryo
  • Mutation

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