Cryopreservation is an important method for the excellent long-term preservation of plant germplasm. This study explores an optimal cryopreservation technology for the embryogenic callus of Fraxinus mandshurica to effectively maintain its genetic stability and morphogenesis potential. The optimal cryopreservation conditions were assessed using the embryogenic callus of F. mandshurica as the material, and the slow cooling method was optimized for its cryopreservation. The results indicated that the preculture of embryogenic callus in 0.4 mol·L−1 sorbitol solution for 20 h at room temperature, followed by its cryoprotection in 7.5% dimethyl sulfoxide solution at 0 °C for 90 min, constituted the optimal material treatment method. The freezing tube was placed in a −80 °C refrigerator for 2 h and then quickly put into liquid nitrogen for frozen storage. During thawing, the cryopreservation tube was taken out from liquid nitrogen, directly placed in a water bath at 40 °C for 2 min, and used for culturing on the woody plant media + 0.1 mg·L−1 6-benzyladenine + 0.15 mg·L−1 2, 4-dichlorophenoxyacetic acid. After cryopreservation using the slow cooling method, the highest survival rate of callus cells was 80.82%. The fresh weight reached 1.93 g after 60-day recovery culture. The regeneration rate and the proliferation coefficient of the callus were 100% and 2.79, respectively. The differentiation rate was 56.83%, and the emergence rate was 23.59%. The results provide a scientific basis for the long-term preservation of F. mandshurica germplasm resources.
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