Optimization of Octoploid Strawberry Regeneration for the Application of Targeted Genome Editing

The octoploid strawberry, Fragaria x ananassa, is cultivated worldwide for its popularity and economic value. Continuous introduction of strawberry varieties with favorable levels of economically important traits is critical to the industry. Many elite varieties have been developed by conventional breeding methods through many generations of phenotypic recurrent selection. This approach is effective for selection of elite genotype from the ancestral germplasm but is very time and resource-intensive. With a targeted genome editing technology (CRISPR/Cas9 system) traits can be modified in a targeted way without disturbing the genetic background of elite strawberry genotypes. However, a bottleneck in application of the technology is a stable procedure of transformation and regeneration for octoploid strawberry. In this report, we describe our initial screening to identify the optimal shoot regeneration conditions from callus for three elite strawberry genotypes bred at the University of Florida. Segments of stolon and petiole from greenhouse-grown plants were used as explants. ‘Florida127’ responded to a basal Murashige and Skoog media containing 1 mg/L of thidiazuron, 0.05 mg/L of 6-benzyladenine and 0.05 mg/L of 2,4-D by inducing somatic embryogenesis on stolons and shoot organogenesis on petioles. The same media also induced somatic embryogenesis of FL12.55-220, but only on petioles. FL 12.121-5 responded at a lower concentration of 2,4-D (0.02 mg/L) in the same media by producing adventitious shoots on stolons. Our finding will make it feasible to apply various biotechnological methods, including targeted genome editing, to the elite strawberry genotypes.