紫参薯的包埋玻璃化超低温保存  

Cryopreservation of Purple Yam(Dioscorea alata) by Encapsulation-vitrification

作　　者：蔡月琴[1] 王艳平[1] 庄君娥 林燕茹[1] 柯美玉 陆銮眉[1] 张泽宏[1] 

机构地区：[1]闽南师范大学生物科学与技术学院,福建漳州363000

出　　处：《西南大学学报（自然科学版）》2016年第9期58-64,共7页Journal of Southwest University(Natural Science Edition)

基　　金：福建省科技计划重点项目(2013N0037);漳州市科技计划项目(ZZ2013052);国家大学生创新训练项目(201410402007)

摘　　要：以紫参薯带芽茎段为材料,采用2%海藻酸钠和0.1mol/L氯化钙凝胶系统制备包埋珠,液氮保存,以茎段细胞活力和包埋珠成活率为指标筛选获得有关参数适宜水平,比较冻后再生苗与常温苗形态及生理指标.结果表明:紫参薯试管苗经4℃锻炼6d后,无菌条件下制备其带芽茎段包埋珠,接种到MS+2 mg/L KT+0.2 mg/L NAA+0.5mol/L蔗糖液体培养基中预培养1d,转入MS+2 mol/L甘油+0.4 mol/L蔗糖溶液中室温下装载40min,在玻璃化保护剂PVS2中脱水40min,更新PVS2后迅即液氮保存,24h后置于37℃水浴中化冻5min,用MS+2mg/L KT+0.2mg/L NAA+0.8mol/L蔗糖溶液洗涤3次(每次10min),然后转入MS+2mg/L KT+0.2mg/L NAA+30g/L蔗糖+1g/L活性炭固体培养基中,暗培养7d后转至光照下培养14d,该包埋珠成活率可达60%.冻后再生苗诸多形态和生理指标与常温苗差异不具有统计学意义.The cryopreservation of purple yam（Dioscorea alata）by Encapsulation vitrification was studied.Stems with axillary buds suspended in Murashige and Skoog（MS）medium supplemented with 2%sodium alginate and 0.4mol/L sucrose were placed in liquid MS medium supplemented with 0.1 mol/L calcium chloride and 0.4mol/L sucrose,and the beads were preserved with liquid nitrogen（LN）.The optimum levels of related parameters were obtained with the stem cell vitality and embedding survival rate as indexes and the comparison of morphological and physiological indexes of regenerated plantlets and normal plantlets was made.The results showed that the plantlets of purple yam were cold-hardened for 6dat 4 ℃,then stems with axillary buds were excised and encapsulated into beads in sterile conditions and then precultured in liquid MS medium supplemented with 2mg/L KT,0.2mg/L NAA and 0.5mol/L sucrose for1 d.The encapsulated beads were loaded with MS medium plus with 2mol/L glycerol and 0.4mol/L sucrose for 40 min at room temperature and then dehydrated with PVS2 for 40min at 0℃.The encapsulated beads were plunged directly into LN for 1day after dehydration and then rapidly thawed in a water bath for5 min at 37 ℃.After warming,the encapsulated beads were rinsed three times with liquid MS medium containing 2mg/L KT,0.2mg/L NAA and 0.8mol/L sucrose for 10 min at 25 ℃,then transferred to regeneration medium supplemented with 2 mg/L KT,0.2 mg/L NAA and 30g/L sucrose plus 0.1g/L activated carbon in the dark for 7dbefore exposure to the light.The survival rate of beads reached to 60%after two weeks of light culture.Plantlets regenerated from cryopreservation had no morphological and physiological variation with the normal plantlets.

关 键 词：紫参薯 带芽茎段 包埋玻璃化 超低温保存 

分 类 号：S632.1[农业科学—蔬菜学] Q945.79[农业科学—园艺学]