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Mutagenesis on a complex mouse genetic background by site-specific nucleases.

Benjamin Davies1,2, Lucy Trelfa3,1, Victoria S Rashbrook3,1

  • 1Wellcome Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford, UK.

Transgenic Research
|August 1, 2024
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Site-specific nucleases efficiently create genetic mutations in complex mouse models for atherosclerosis research. This method bypasses lengthy backcrossing, accelerating gene function studies and offering animal welfare benefits.

Keywords:
Abcg13RsAtherosclerosisRegressionSite directed mutagenesis

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Area of Science:

  • Genetics
  • Cardiovascular Research
  • Preclinical Models

Background:

  • Complex mouse models are crucial for preclinical research but require time-consuming backcrossing.
  • Developing new genetic modifications on existing complex backgrounds is challenging.

Purpose of the Study:

  • To evaluate the efficacy of site-specific nucleases for generating mutations in a complex genetic background.
  • To investigate the role of ATP binding cassette transporter G1 (ABCG1) in atherosclerosis regression using this method.

Main Methods:

  • Utilized the REVERSA mouse model for atherosclerosis, which has four genetic alterations.
  • Microinjected site-specific nucleases into zygotes from the REVERSA model to create ABCG1 knockouts.
  • Assessed cholesterol efflux in bone marrow-derived macrophages and atherosclerosis regression in the aortic root and arch.

Main Results:

  • Successfully generated two independent ABCG1 knockout lines on the REVERSA background.
  • Observed a significant reduction in cholesterol efflux to HDL in macrophages from knockout mice.
  • Found no impact of Abcg1 loss on atherosclerosis regression in the aortic root or arch.

Conclusions:

  • Site-specific nucleases are effective for creating genetic modifications directly on complex disease backgrounds.
  • This approach facilitates gene function exploration without extensive backcrossing, aligning with the 3Rs principles.
  • ABCG1 does not play a significant role in atherosclerosis regression in this model.