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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 purple ginseng tuber bud section was used as material, and 2% sodium alginate and 0.1 mol / L calcium chloride gel system were used to prepare the embedded beads. The cells were stored in liquid nitrogen and the viability of the stem cells and the survival rate of the embedded beads were taken as indexes The appropriate parameters were obtained by screening, and the morphological and physiological indices of seedlings were compared between the frozen seedlings and the normal seedlings.The results showed that the seedlings of the test-tube seedlings were cultured under 4 ℃ for 6 days under aseptic conditions, The cells were preincubated for 1 day in MS + 2 mg / L KT + 0.2 mg / L NAA + 0.5 mol / L sucrose liquid medium and then transferred to MS + 2 mol / L glycerol + 0.4 mol / L sucrose solution for 40 min at room temperature, PVS2 was vitrified for 40min, and after PVS2 was renewed, it was quickly stored in liquid nitrogen. After 24h, it was thawed in water bath at 37 ° C for 5min and washed with MS + 2mg / L KT + 0.2mg / L NAA + 0.8mol / L sucrose solution (10 min each), and then transferred to MS + 2mg / L KT + 0.2mg / L NAA + 30g / L sucrose + 1g / L activated carbon solid medium for 7 days and then transferred to light for 14 days The survival rate of embedded beads can reach 60% .No significant difference was found in the number of morphological and physiological indexes of regenerated seedlings after cold-thaw.