Rapid non-GMO CRISPR genome editing in plant seeds via optimised lipofection delivery

Overview

This solution offers a rapid, non-GMO CRISPR-Cas9 genome editing platform for plant seeds that bypasses the need for transgenic intermediates, zygote manipulation, or in vitro fertilisation. By delivering the sgRNA-Cas9 ribonucleoprotein (RNP) complex directly into seeds, the method generates inheritable edits and observable mutant phenotypes in the F0 generation, dramatically shortening the timeline for functional genomics and crop trait development. The approach is being optimised through systematic comparison of leading lipofection reagents to maximise editing efficiency, with the goal of creating a universal editing tool applicable to any crop with a sequenced genome, including potential field deployment.

Technical specifications

Key features:

• Non-transgenic editing: Delivers pre-assembled Cas9-sgRNA RNP complexes directly into seeds, avoiding foreign DNA integration
• F0 phenotyping: Mutant phenotypes manifest in the first generation, eliminating multi-generation breeding cycles
• Inheritable edits: Confirmed germline transmission of edits and phenotypes to the next generation across three validated gene knockouts
• Lipofection optimisation: Systematic evaluation of leading lipid-based transfection reagents, including those used for whole-organism delivery, to improve RNP uptake efficiency
• Broad species applicability: Designed for transfer from the Arabidopsis thaliana model to both monocot and dicot crop species
• Field-deployable potential: Absence of transgenic elements supports possible in-field application subject to regulatory frameworks

Technology readiness level

The method has been validated through successful knockout of three genes in Arabidopsis thaliana, with both edits and phenotypes confirmed as heritable. Proof-of-concept data demonstrates functional RNP delivery and editing in seeds. The current project focuses on two work packages: (1) optimising delivery efficiency by comparing leading lipofection reagents against the existing naked delivery approach through sequencing and phenotyping of large edited seedling batches, and (2) extending the optimised method to monocot and dicot crop seedlings in collaboration with a crop-science partner. The technology is at an early-to-mid stage of development, with established proof-of-concept in a model organism and ongoing translational work toward agricultural crops.