Thidiazuron Enhances Strawberry Shoot Multiplication by Regulating Hormone Signal Transduction Pathways

Tissue culture-based rapid propagation is critical for genetic improvement and virus-free production of strawberries (Fragaria × ananassa). This study evaluated the optimal concentration of thidiazuron (TDZ) for shoot multiplication and explored the underlying molecular mechanisms. Strawberry explants were treated with TDZ at concentrations of 0, 0.025, 0.05, 0.1, and 0.4 mg·L^-1 in vitro, and growth, physiological changes, and transcriptomic profiles were analyzed after four weeks. The results identified 0.05 mg·L^-1 TDZ as the most effective concentration for shoot proliferation, yielding a significant increase in leaf number. However, TDZ application inhibited plant height and reduced chlorophyll, carotenoid, and soluble sugar contents. Physiological analyses revealed that TDZ decreased endogenous cytokinin levels while elevating auxin concentrations. Transcriptomic analysis showed that TDZ suppressed cytokinin biosynthesis and up-regulated cytokinin oxidase expression, thereby modulating hormone homeostasis. Additionally, TDZ enhanced the cytokinin signaling pathway, which is crucial for cell division and shoot initiation, and influenced auxin, gibberellin, and brassinosteroid pathways to regulate differentiation. These findings suggest that TDZ promotes strawberry shoot multiplication primarily through hormone signal transduction, providing insights for optimizing tissue culture protocols.