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An engineered prime editor with enhanced editing efficiency in plants.

Yuan Zong1, Yijing Liu1,2, Chenxiao Xue1,2

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Optimized prime editors significantly enhance genome editing efficiency. Modifications to reverse transcriptase and nucleocapsid protein improve prime editing in plant cells and crops.

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Area of Science:

  • Molecular Biology
  • Plant Biotechnology
  • Genome Editing

Background:

  • Prime editing is a powerful genome-editing tool with broad applications.
  • However, its practical use is limited by low editing efficiency in many contexts.

Purpose of the Study:

  • To engineer and optimize prime editors for substantially improved editing efficiency.
  • To enhance the precision and efficacy of prime editing in plant systems.

Main Methods:

  • Engineered the Moloney-murine leukemia virus reverse transcriptase by removing its ribonuclease H domain.
  • Incorporated a viral nucleocapsid protein with nucleic acid chaperone activity.
  • Combined modifications in an engineered plant prime editor (ePPE) and tested on endogenous sites and in rice.

Main Results:

  • Individual modifications improved prime editing efficiency by 1.8-3.4-fold in plant cells.
  • The combined ePPE synergistically enhanced base substitutions, deletions, and insertions by an average of 5.8-fold compared to the original prime editor (PPE).
  • Achieved 11.3% editing frequency in rice for herbicide tolerance using ePPE, compared to 2.1% with PPE, with no significant increase in byproducts or off-target editing.

Conclusions:

  • The engineered plant prime editor (ePPE) demonstrates significantly enhanced efficiency and specificity.
  • ePPE holds great promise for accelerating crop improvement and other genome-editing applications in plants.
  • Further improvements were achieved by combining ePPE with engineered prime editing guide RNAs.