Selection of the high efficient sgRNA for CRISPR-Cas9 to edit herbicide related genes, PDS, ALS, and EPSPS in tomato

Abstract Herbicide resistance is one of the main crop traits that improve farming methods and productivity. CRISPR-Cas9 can be applied to development herbicide-resistant crops based on a target site mechanism, by editing genes encoding herbicide binding proteins. The sgRNAs capable herbicides, pds (phytoene desaturase), ALS (acetolactate synthase), EPSPS (5-Enolpyruvylshikimate-3-phosphate were designed use with system in tomato ( Solanum lycopersicum cv. Micro-Tom). efficiency was tested using Agrobacterium mediated transient expression cotyledons. One sgRNA for PDS had no significant efficiency. However, three different efficiency, them, ALS2-P sgRNA, showed over 0.8% average cotyledon genome. maximum around 1.3%. An 0.4% average. testing provided confidence sites could achieved transformation process. We confirmed 19 independent transgenic tomatoes successfully edited ALS2_P or ALS1_W two them base deletion mutations, which are expected have altered resistance. In this study, we demonstrated usefulness performing an test before transformation, valuable tool breeding crops.