This study aims to develop an efficient tissue culture propagation system for Eucalyptus maidenii and identify high- performance genotypes, thereby establishing a theoretical and technical foundation for large-scale seedling production.

Seeds from elite E. maidenii families were used as explants. Using full factorial and orthogonal experimental designs, the optimal culture conditions were determined for primary culture, subculture proliferation, adventitious root induction, and hardening. The selected optimal proliferation and rooting media were then applied to axenic seedlings of different genotypes to screen genotypes that exhibited both high proliferation efficiency and robust rooting competence.

The optimal conditions for the primary culture and subculture of sterile stem segments of E. maidenii are MS medium supplemented with 0.5 mg/L 6-BA, 0.6 mg/L NAA, 20 g/L sucrose, and 4.5 g/L agar, resulting in a 100% axillary bud induction rate, a 10.40-fold monthly proliferation rate, a plantlet height of 1.14 cm, and robust shoot clusters. Adventitious root induction is optimally achieved on 1/2 DCR medium containing 0.6 mg/L IBA, 0.3 mg/L NAA, 1.0 mg/L ABT1, 20 g/L sucrose, 5 g/L agar, and 0.16 g/L activated charcoal, with a 86.67% rooting rate after 20 days. A 1:1:1 (v/v/v) mixture of humus soil, vermiculite, and peat soil provides the highest transplantation success, with a 76.67% survival rate. A total of 118 E. maidenii genotypes were evaluated for proliferation and rooting capacity, from which one superior genotype (G116) with both high proliferation efficiency and strong rooting ability was selected. After 60 days of culture, G116 exhibits a 11.76-fold proliferation rate, reaches 1.65 cm in height, and develops robust shoot clusters. This genotype also demonstrates a 93.33% rooting rate within 20 days of culture.

The research successfully developed an efficient in vitro propagation system for E. maidenii and selected one genotype exhibiting superior propagation traits. This methodology enhanced seedling production efficiency.