Efficient generation of stable, heritable gene edits in wheat using CRISPR/Cas9

Area of Science:

• Plant Biotechnology
• Molecular Genetics
• Agricultural Science

Background:

• CRISPR/Cas9 systems are powerful tools for crop research.
• They enable gene function knockout and precise allele adjustment in complex plant genomes.

Purpose of the Study:

• Compare CRISPR/Cas9 gene editing efficiency and stability in hexaploid wheat (Triticum aestivum) and diploid barley (Hordeum vulgare).
• Investigate single genome and tri-genome targeting strategies in wheat.

Main Methods:

• Stable Agrobacterium-mediated transformation for gene editing in wheat.
• Comparative analysis of gene editing outcomes in wheat and barley across generations.

Main Results:

• Achieved high gene editing efficiency in wheat (11-17% for single genome, 5% for tri-genome guides).
• Wheat gene editing resulted in predominantly heterozygous, Mendelian-inherited edits with no off-target effects.
• Wheat produced more stable, heritable edits compared to barley, which showed continued somatic editing.

Conclusions:

• Stable, edited, transgene-free wheat lines can be obtained in 36 weeks within two generations.
• Targeted mutagenesis of individual homeologues in the wheat genome is feasible without off-target mutations or extensive crossing.