Abstract
Polyploid crops, such as wheat, potato, and alfalfa, present unique challenges for genome editing due to their redundant genomes and complex genetics. CRISPR-Cas9 base editing offers a promising approach for precise nucleotide modifications without double-strand breaks, enabling targeted trait improvement. This study evaluates the efficiency and specificity of cytosine and adenine base editors in three polyploid crop species: hexaploid wheat (Triticum aestivum), tetraploid potato (Solanum tuberosum), and autotetraploid alfalfa (Medicago sativa). Using ribonucleoprotein (RNP) delivery and protoplast regeneration, we targeted six genes associated with agronomic traits (e.g., ALS for herbicide resistance, GBSS for starch quality). Base editing efficiencies ranged from 2.1% to 18.7% across species and targets, with cytosine base editors showing higher activity in GC-rich contexts. Off-target analysis via targeted deep sequencing revealed minimal off-target edits (
Keywords
CRISPR-Cas9, base editing, polyploid crops, precise gene modification, wheat, potato, alfalfa, genome editing