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Efficient prime editing in two-cell mouse embryos using PEmbryo.

Rebecca P Kim-Yip1, Ryan McNulty2, Bradley Joyce1

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Summary
This summary is machine-generated.

Transiently inhibiting DNA mismatch repair boosts prime editing efficiency in mouse embryos. This advance allows for same-generation phenotyping with minimal errors, though it may increase off-target mutations.

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Area of Science:

  • Genetics
  • Molecular Biology
  • Developmental Biology

Background:

  • Prime editing is a powerful gene editing technology.
  • Efficient in vivo gene editing in mammalian embryos is crucial for genetic studies.
  • DNA mismatch repair (MMR) can interfere with precise gene editing outcomes.

Purpose of the Study:

  • To investigate the impact of transient MMR inhibition on prime editing efficiency in mouse embryos.
  • To assess the precision and potential off-target effects of prime editing under MMR-inhibited conditions.
  • To enable same-generation phenotyping of genetically modified founder mice.

Main Methods:

  • Transiently inhibited DNA mismatch repair (MMR) in mouse embryos during a specific developmental window.
  • Utilized prime editing to introduce precise genetic modifications.
  • Performed whole-genome sequencing to evaluate on-target and off-target editing events.
  • Assessed the phenotype of founder mice generated from edited embryos.

Main Results:

  • Achieved highly efficient prime editing with an average precise edit frequency of 58%.
  • Observed a low on-target error frequency of 0.5% across multiple edits and sites.
  • Demonstrated successful same-generation phenotyping of founder mice.
  • Identified an increase in off-target insertions/deletions (indels) at low-complexity genomic regions due to MMR inhibition, without causing apparent phenotypes in mice.

Conclusions:

  • Transient MMR inhibition is a viable strategy to enhance prime editing efficiency in mouse embryos.
  • This method facilitates rapid genetic analysis and phenotyping in founder animals.
  • While effective, MMR inhibition necessitates careful evaluation of potential genomic off-target effects.