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Related Concept Videos

In-vitro Mutagenesis01:16

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To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.
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Efficient Genome Editing of Mice by CRISPR Electroporation of Zygotes
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A universal method for generating knockout mice in multiple genetic backgrounds using zygote electroporation.

Tomohiro Tamari1,2,3, Yoshihisa Ikeda1,4, Kento Morimoto5,6

  • 1Model Generation & Breeding Service, The Jackson Laboratory Japan, Inc., 955 Kamibayashi, Ishioka, Ibaraki 315-0138, Japan.

Biology Open
|August 25, 2023
PubMed
Summary

This study shows a new, efficient method for creating genetically engineered knockout mice in various strains using CRISPR-Cas9 zygote electroporation. This advance aids in studying gene function and diseases across different mouse models.

Keywords:
In vitro fertilisationElectroporationGenome editingKnockoutLong-read sequencingMouse

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Area of Science:

  • Genetics and Genomics
  • Mammalian Genetics
  • Molecular Biology

Background:

  • Genetically engineered mouse models are crucial for studying gene function and disease.
  • Genome editing technologies, like CRISPR-Cas9, enable rapid generation of these models.
  • Zygote electroporation offers a cost-effective and labor-efficient method for gene editing.

Purpose of the Study:

  • To demonstrate the effectiveness of a generalized zygote electroporation method for generating knockout mice across multiple inbred strains.
  • To establish a versatile approach for creating genetically modified mouse models for diverse research needs.

Main Methods:

  • Utilized in vitro fertilization combined with zygote electroporation to introduce CRISPR-Cas9 complex.
  • Generated founder knockout mice targeting specific alleles in eight common inbred mouse strains.
  • Employed long-read sequencing for detailed analysis of intended and unintended mutant alleles.

Main Results:

  • Successfully generated knockout mice founders in eight different inbred strains.
  • Identified variations in allele frequencies (intended vs. unintended) among strains via sequencing.
  • Confirmed successful germline transmission of the generated knockout alleles.

Conclusions:

  • The generalized zygote electroporation method is effective for creating knockout mice in multiple inbred strains.
  • This approach facilitates the establishment of mutant mice targeting the same locus across various strains for comparative phenotyping.
  • Contributes to advancing reverse genetics strategies and human disease research through versatile mouse model generation.