Plant, cell & environmentSep 9, 2025

Gene Editing in Plants Without Adding Foreign DNA: Progress, Challenges, and Future for Trees

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗

Updated Jan 3, 2026

Abstract

CRISPR ribonucleoprotein (RNP)-mediated genome editing may offer a transgene-free platform for precise genetic modification in various species, including rice, wheat, and grapevine.

Woody plants, such as bamboo, face distinct challenges in RNP-mediated editing due to inefficient delivery and regeneration difficulties.
Complexities in woody species arise from factors like intricate cell wall architecture and widespread recalcitrant genotypes.
RNP editing utilizes protein-guide RNA complexes without integrating foreign DNA, potentially simplifying regulatory approval.
Current RNP delivery methods primarily include PEG-mediated protoplast transfection and particle bombardment, though alternative strategies exist.
Promising alternative delivery methods, such as lipofection and nanoparticle-based systems, remain largely untested in woody species.
Future research may focus on developing tree-specific RNP delivery systems and regeneration protocols to enhance editing efficiency.

Simplified

CRISPR ribonucleoprotein (RNP)-mediated genome editing offers a transgene-free platform for precise genetic modification in diverse herbaceous and tree species, including rice, wheat, apple, poplar, oil palm, rubber tree and grapevine. However, its application in woody plants faces distinct challenges, notably inefficient delivery and regeneration difficulties, particularly in species such as bamboo. While some of these issues also occur in herbaceous plants, they are often significantly more complex in woody species due to factors such as intricate cell wall architecture, widespread recalcitrant genotypes and inherent limitations of current delivery platforms. This review presents the first in-depth, critical re-evaluation of recent advancements in RNP-mediated editing in woody plants, highlighting these obstacles that warrant focused attention. Unlike plasmid-based CRISPR systems, RNP editing utilises Cas9/Cas12a protein-guide RNA complexes without integrating foreign DNA. This enables a DNA-free editing strategy that simplifies regulatory approval and minimises off-target effects due to the transient presence and rapid degradation of RNPs within plant cells. While PEG-mediated protoplast transfection and particle bombardment remain the primary reported methods for RNP delivery in trees, we evaluate promising alternative strategies such as lipofection, electroporation, cell-penetrating peptides and nanoparticle-based systems for targeted RNP delivery. Despite their promise, these advanced methods remain largely untested in woody species. Finally, we outline future research directions, including the development of tree-specific RNP delivery systems and regeneration protocols to enhance efficiency and minimise cytotoxicity. These innovations are essential for unlocking the full potential of RNP-mediated genome editing in long-lived tree species. This review provides a focused and timely roadmap for expanding the application of RNP technology across diverse woody plants.

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Gene Editing in Plants Without Adding Foreign DNA: Progress, Challenges, and Future for Trees

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