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PLoS One. 2018 May 3;13(5):e0196891. doi: 10.1371/journal.pone.0196891. eCollection 2018.

An optimized electroporation approach for efficient CRISPR/Cas9 genome editing in murine zygotes.

Author information

1
Cologne Excellence Cluster for Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany.
2
in vivo Research Facility, Medical Faculty, University of Cologne, Cologne, Germany.
3
Department II of Internal Medicine and Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany.
4
Systems Biology of Aging Cologne (Sybacol), University of Cologne, Cologne, Germany.

Abstract

Electroporation of zygotes represents a rapid alternative to the elaborate pronuclear injection procedure for CRISPR/Cas9-mediated genome editing in mice. However, current protocols for electroporation either require the investment in specialized electroporators or corrosive pre-treatment of zygotes which compromises embryo viability. Here, we describe an easily adaptable approach for the introduction of specific mutations in C57BL/6 mice by electroporation of intact zygotes using a common electroporator with synthetic CRISPR/Cas9 components and minimal technical requirement. Direct comparison to conventional pronuclear injection demonstrates significantly reduced physical damage and thus improved embryo development with successful genome editing in up to 100% of living offspring. Hence, our novel approach for Easy Electroporation of Zygotes (EEZy) allows highly efficient generation of CRISPR/Cas9 transgenic mice while reducing the numbers of animals required.

PMID:
29723268
PMCID:
PMC5933690
DOI:
10.1371/journal.pone.0196891
[Indexed for MEDLINE]
Free PMC Article

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