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<p>Genome editing, in particular the CRISPR/Cas9 system, is widely used to generate new animal models. However, the generation of mutations, such as conditional knock-out or knock-in, can remain complex and inefficient, in particular because of the difficulty to deliver the donor DNA (single or double stranded) into the nucleus of fertilized oocytes. The use of recombinant adeno-associated viruses (rAAV) as donor DNA is a rapidly developing approach that promises to improve the efficiency of creation of animal models. In this mini-review, we explore the progress and challenges of using CRISPR/Cas9 in combination with rAAV for precise genome editing. We will summarise the current knowledge of rAAV transduction, data on its use in rodent embryos in combination with CRISPR/Cas9 to easily generate sequence replacements or insertions, the limitations of rAAV and the unexpected events observed to date, and the protocol optimisations already in place to facilitate its use in the generation of animal models.</p>

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Genome engineering with Cas9 and AAV repair templates, successes and pitfalls

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年1月13日

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  • M.C. Birling,
  • Y. Hérault,
  • G. Pavlovic

摘要

Genome editing, in particular the CRISPR/Cas9 system, is widely used to generate new animal models. However, the generation of mutations, such as conditional knock-out or knock-in, can remain complex and inefficient, in particular because of the difficulty to deliver the donor DNA (single or double stranded) into the nucleus of fertilized oocytes. The use of recombinant adeno-associated viruses (rAAV) as donor DNA is a rapidly developing approach that promises to improve the efficiency of creation of animal models. In this mini-review, we explore the progress and challenges of using CRISPR/Cas9 in combination with rAAV for precise genome editing. We will summarise the current knowledge of rAAV transduction, data on its use in rodent embryos in combination with CRISPR/Cas9 to easily generate sequence replacements or insertions, the limitations of rAAV and the unexpected events observed to date, and the protocol optimisations already in place to facilitate its use in the generation of animal models.