摘要
对柿 (DiospyroskakiThunb .)休眠芽茎尖进行了玻璃化法超低温保存及植株再生研究 ,结果表明 ,1.5～2 .0mm茎尖在含有 0 .3、0 .5和 0 .7mol·L-1蔗糖的改良MS培养基 (KNO3 和NH4NO3 减半 )上预培养各 1d ,室温(2 0℃ )下装载液 (2mol·L-1甘油 ,0 .4mol·L-1蔗糖 )停留 2 0min ,0℃下玻璃化液 (PVS2 或PVS3 )处理 30～ 4 0min ,投入液氮保存 1d后 ,4 0℃水浴化冻 1min ,含蔗糖 1.2mol·L-1的改良MS培养基洗涤 2 0min ,接种于含 0 .2mg·L-1CPPU、1mg·L-1BA、0 .0 5mg·L-1IAA、5 0 0mg·L-1可溶性PVP、30g·L-1蔗糖和 7g·L-1琼脂的改良MS培养基 (再生培养基 )表面的滤纸上 ,暗处理 1d后转移到新鲜的上述再生培养基中 ,暗培养 1周后转到正常光下 ,成活率高达 70 .2 % ,再生植株生长和分化正常。本研究结果为柿种质的长期保存提供了新途径。
A procedure on cryopreservation of dormant shoot tips of persimmon(Diospyros kaki Thunb.)by vitrification was studied. Shoot tips, 1.5-2.0 mm in length, were precultured on the modified MS medium with nitrates reduced to half strength (MS(1/2N)) supplemented with sucrose at concentrations of 0.3, 0.5 and 0.7 mol·L -1 for 3 days, loaded with the MS(1/2N)medium supplemented with 2 mol·L -1 glycerol and 0.4 mol·L -1 sucrose for 20 min at 20℃, and incubated in PVS 2 or PVS 3 for 30-40 min at 0℃ prior to a direct plunge into liquid nitrogen (LN). After rapid thawing in water at 40℃, the shoot tips were rinsed in the MS(1/2N)medium supplemented with 1.2 mol·L -1 sucrose for 20 min,plated on the filter paper sustained by the MS(1/2N)regeneration medium supplemented with 0.2 mg·L -1 CPPU,1 mg·L -1 BA,0.05 mg·L -1 IAA, 500 mg·L -1 soluble PVP, 30 g·L -1 sucrose and 7 g·L -1 agar for 1d in dark and subcultured on the above regeneration medium for one week in dark prior to exposure to the light. The survival of shoot tips was up to 70.2% after 6 weeks and they grew and differentiated normally. Thus, this simple and reliable vitrification protocol provided a new strategy for long term preservation of persimmon germplasm.
出处
《中国农业科学》
CAS
CSCD
北大核心
2003年第5期553-556,共4页
Scientia Agricultura Sinica
