Engineering plant architecture in the ornamental species Eustoma grandiflorum by knockout of strigolactone...

The targeted mutation of the strigolactone biosynthetic gene, CCD8, through genome-editing CRISPR-Cas9 induces dwarfism and enhanced branching in Eustoma grandiflorum. In ornamental plants, modifying shoot architecture is a major breeding objective for diverse applications; however, achieving this goal using conventional techniques remains challenging. Strigolactones are plant hormones that inhibit shoot branching. We aimed to generate a strigolactone-deficient mutant of Eustoma grandiflorum using CRISPR-Cas9 genome editing. Four guide RNAs targeting CAROTENOID CLEAVAGE DIOXYGENASE 8 (EgCCD8) were designed, and transgenic E. grandiflorum plants were produced. Three independent lines carried deletions or insertions in all EgCCD8 alleles, indicating high editing efficiency. Two lines exhibited increased branching and pronounced dwarfism. In one of these lines, plant height in T₁ null segregants was reduced to 47% of wild-type (WT) levels. Branch number increased 5.0-fold compared with the WT, and flower bud formation increased 1.7-fold alongside enhanced branching. These findings demonstrate that the targeted mutation of CCD8 through genome editing induces dwarfism and enhanced branching without compromising other ornamental traits.