摘要
主要探讨蔗糖浓度、装载时间和PVS2处理时间对部分中国甜柿种质超低温保存后成活率的影响。结果表明,长约1 mm的柿休眠芽茎尖用海藻酸钙包埋后,在含有0.3 molL-1蔗糖的改良MS培养基(KNO3和NH4NO3减半)上预培养1 d,室温(25℃)下PVS2处理2 h后快速投入液氮(LN),1 h后取出,在40℃水浴中化冻1～2 min,用含有1.2 molL-1蔗糖的改良MS培养液洗涤20 min,接种于含ZT(玉米素)2.2 mgL-1、PVP(聚乙烯吡咯烷酮)600 mgL-1的改良MS培养基,暗培养3～5 d,转移至正常光下,最高成活率接近100%;再生植株生长和分化正常。
Shoot tips excised from dormant axillary buds of non-astringent types of Chinese persimmon(Diospyros kaki Thunb.), which is originated from China, had been cryopreserved by encapsulation-vitrification for the first time. In the experiment, three factors, namely sucrose concentration, loading time and exposure time to PVS2,were considered and a good protocol was achieved. Excised shoot tips were encapsulated into alginated-gel beads. Then alginate-coated shoot tips were precultured on a modified MS basal medium supplemented with sucrose at 0.3, 0.5, 0.7, 1.0 molL-1 for one day. After preculture, encapsulated shoot tips were loaded with a mixture of 2 mol稬-1 glycerol plus 0.4 mol稬-1sucrose for 20 min. Following loading, encapsulated shoot tips were dehydrated with a highly concentrated vitrification solution (PVS2 solution)for 2 h at 25℃, and then plunged directly into liquid nitrogen. After rapid warming in water for about one to two min at 40℃, the shoot tips were rinsed with the modified MS medium containing 1.2 molL-1 sucrose for 20 min, and then cultured on the modified medium with 2.2 mg稬-1ZT and 600 mg稬-1 PVP in dark prior to exposure to the light. The best survival rate of encapsulation vitrified shoot tips amounted to nearly 100% and no abnormality was observed. Thus the protocol using dormant shoot tips appeared to be promising as a routine method for the long-term conservation of Diospyros spp. germplasm.
出处
《中国农业科学》
CAS
CSCD
北大核心
2004年第12期2019-2022,共4页
Scientia Agricultura Sinica
基金
国家自然科学基金资助项目(30270925)
