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Breeding by Design for Functional Rice with Genome Editing Technologies
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Efficient prime editors for heritable multiplex precision genome editing in soybean.

Fei Su1, Yinghui Dong1, Rui Guo1

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Summary

We developed GmPEplus, an optimized prime editing (PE) system for soybean, significantly boosting editing efficiency. This system enables multiplex gene editing, offering powerful tools for soybean genetic improvement.

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Area of Science:

  • Plant Biotechnology
  • Molecular Biology
  • Genetics

Background:

  • Prime editing (PE) is a powerful genome editing technology.
  • Low efficiency of PE in dicot plants, including soybean, limits its application.
  • Development of enhanced PE systems is crucial for crop improvement.

Purpose of the Study:

  • To develop an optimized and highly efficient prime editing system for soybean.
  • To establish a multiplex prime editing system for simultaneous gene editing in soybean.
  • To provide versatile tools for precise and heritable genome editing in soybean breeding.

Main Methods:

  • Engineered the prime editor (PE) by modifying Cas9 and reverse transcriptase domains and incorporating a viral protein.
  • Co-expressed a dominant-negative GmMLH1 allele to enhance editing.
  • Utilized a tRNA processing system and AtU6 promoter for sgRNA expression optimization.
  • Developed a Csy4-mediated multiplex PE system (CMMPE) for simultaneous editing.

Main Results:

  • The optimized system, GmPEplus, achieved up to 81.3% editing efficiency in stable transgenic soybean lines.
  • Optimizing sgRNA expression boosted editing efficiency by 13.1-fold.
  • CMMPE enabled simultaneous editing of 2-12 genes in soybean hairy roots and up to 3 genes in stable lines.

Conclusions:

  • GmPEplus significantly enhances prime editing efficiency and versatility in soybean.
  • CMMPE allows for multiplex and simultaneous genome editing in soybean.
  • These advanced editing tools are valuable for soybean breeding and genetic research.