Paul H, Daigny G, Sangwan-Norreel B S
Faculté des Sciences, Laboratoire Androgenèse et Biotechnologie, 33 rue Saint Leu, 80039 Amiens Cedex 01, France Fax: +33-0322-827612 e-mail
Plant Cell Rep. 2000 Jul;19(8):768-774. doi: 10.1007/s002990000195.
Axillary shoot tips of apple cv. Golden Delicious isolated from shoot cultures were successfully cryopreserved using the encapsulation-dehydration technique. After encapsulation in alginate gel, embedded shoot tips were dehydrated by exposure to a sterile air flow before being frozen in liquid nitrogen and subsequent slow thawing. A preculture on modified MS medium containing 0.75 M sucrose followed by 6 h of dehydration (21% residual water) led to the highest shoot regrowth of frozen, coated shoot tips (83.7%). Among the sugars tested, sucrose and sorbitol presented the best cryoprotective effect. Four other scion apple varieties and rootstocks were also successfully cryopreserved. Axillary shoot tips of five apple (Malus×domestica Borkh.) scion and rootstock cultivars were cryopreserved using the encapsulation-vitrification technique. Using a one-step freezing method, we successfully cryopreserved axillary shoot tips without the requirement of a cold hardening pretreatment of the shoot cultures. Cryopreserved shoot tips treated with aqueous cryoprotective mixture IV containing 180% (w/v) sucrose and 120% (v/v) ethylene glycol showed the highest shoot regrowth rates, which varied from 64% to 77%, depending on the cultivar.
采用包埋脱水技术成功对从苹果茎尖培养物中分离得到的苹果品种金冠的腋芽茎尖进行了超低温保存。在包埋于藻酸盐凝胶后，将包埋的茎尖置于无菌气流中脱水，然后在液氮中冷冻并随后缓慢解冻。在含有0.75 M蔗糖的改良MS培养基上预培养，接着脱水6小时（残留水分21%），可使冷冻包被茎尖的茎尖再生率最高（83.7%）。在所测试的糖类中，蔗糖和山梨醇具有最佳的冷冻保护效果。其他四个苹果接穗品种和砧木也成功进行了超低温保存。采用包埋玻璃化法对五个苹果（苹果属×家养苹果博尔克）接穗和砧木品种的腋芽茎尖进行了超低温保存。使用一步冷冻法，我们成功地对腋芽茎尖进行了超低温保存，而无需对茎尖培养物进行低温驯化预处理。用含有180%（w/v）蔗糖和120%（v/v）乙二醇的水性冷冻保护混合物IV处理的超低温保存茎尖显示出最高的茎尖再生率，根据品种不同，再生率在64%至77%之间。
