Droplet-vitrification for cassava shoot tip cryopreservation: protocol optimization and histological observation for rapid shoot tip recovery
文献类型： 外文期刊
作者： Wang, Min-Rui 1 ; Miao, Xiu-Qing 4 ; Fu, Yun-Liu 1 ; Chen, Lang-Xin 1 ; Wang, Xiao-Bing 1 ; Yang, Qing-Quan 1 ; Xu, Li 1 ; Li, Zhiying 1 ;
作者机构： 1.Chinese Acad Trop Agr Sci, Trop Crop Genet Resources Inst, Haikou 571101, Hainan, Peoples R China
2.Hainan Prov Key Lab Trop Crops Germplasm Resources, Hainan Vitro Gene Bank Trop Crops, Natl Gene Bank Trop Crops, Danzhou 571737, Hainan, Peoples R China
3.Key Lab Minist Agr Germplasm Resources Conservat &, Haikou 571101, Peoples R China
4.Hainan Univ, Coll Trop Agr & Forestry, Haikou 570228, Peoples R China
关键词： Cassava; Cryopreservation; Droplet-vitrification; Histological study
期刊名称：PLANT CELL TISSUE AND ORGAN CULTURE （ 影响因子：2.4； 五年影响因子：2.6 ）
ISSN： 0167-6857
年卷期： 2025 年 162 卷 2 期
页码：
收录情况： SCI
摘要： Shoot tip cryopreservation is an important strategy for the long-term conservation of vegetatively propagated plant species, but requires protocol optimization when applied to new genera or species. Droplet-vitrification (Dr-vi) has emerged as an efficient cryopreservation protocol for a wide range of plant genus but its optimization for cassava (Manihot esculenta Crantz) remains underexplored. In this study, key steps of the Dr-vi protocol for cassava shoot tip cryopreservation were systematically optimized using MS culture media devoid of plant growth regulators. Our findings indicate that cassava shoot tips were not tolerant to long sucrose preculture, while PVS2 treatment on ice for 40-60 min proved to be an optimal cryoprotective treatment compared to room temperature treatments. The cryopreserved shoot tips could be rewarmed with 0.6-1.2 M sucrose, and direct recovery on the solid regrowth medium was recommended. Genotype response assays revealed that the optimized Dr-vi enabled shoot tip regrowth in most tested genotypes with varied efficiencies. To investigate the basis underlying the rapid regrowth observed in certain genotypes, comprehensive histological studies were implemented, revealing well preserved apical meristem, viable procambium and epidermis cells in shoot tip after cryopreservation. Since root regeneration is crucial for subsequent rapid shoot elongation, histological observation further noted the regeneration of adventitious root meristem at 7-day post thaw culture (PTC) and well-defined roots by 14-day PTC. Our study highlights the critical role of protocol optimization in cryopreservation research, providing valuable insights to enhance cassava cryobanking and the understanding of post-cryopreservation plantlet regrowth.
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