摘要
以马铃薯试管苗为试材,对其茎尖小滴玻璃化法超低温保存的影响因素进行了研究,并对再生植株进行了遗传稳定性检测。结果表明,马铃薯茎尖依次在含有0.3mol·L-1和0.5mol·L-1蔗糖的液体MS培养基中预培养各1d后,在0℃下PVS2处理30min,转到铝箔条上PVS2小滴上(约15μL),将粘有茎尖的铝箔条在液氮里蘸一下,然后直接装入盛满液氮的冷冻管中,投入液氮至少保持1h。室温下用含有1.2mol·L-1蔗糖的MS液体培养基解冻并洗涤30min后,接种到MS+0.5mg·L-1Zeatin+0.1mg·L-1NAA+1.0mg·L-1GA3恢复培养基上,存活率和再生率最高达79.91%和62.52%。通过SSR分子标记检测,再生植株的遗传稳定性没有发生改变。
Shoot tips from axillary buds of in vitro potato（Solanum tuberosum L.）were successfully cryopreserved by droplet vitrification.The optimum cryopreservation procedures were as follows.Shoot tips excised from 2–3 month-old plantlets were precultured on liquid MS medium supplemented with 0.3 mol · L^-1 and 0.5 mol · L^-1 sucrose for 1 day each and then dehydrated with PVS2 for 30 min at 0 ℃.Five shoot tips were transferred to approximately 15 μL droplets of PVS2 solution on thin strips of sterile aluminum foil.The aluminum foil strips were folded to enclose the shoot tips.The foil envelope was then carefully immersed into liquid nitrogen（LN）using fine forceps.After immersion the strips were quickly transferred to 2 mL cryotubes and immediately plunged into LN and maintained for 1 h at least.The shoot tips in the foil strips were then rapidly heated by immersion twice at 15 min intervals into 10 mL 1.2 mol · L^-1 sucrose MS medium at room temperature.The shoot tips were then unloaded from the foil, rinsing the PVS2 from the shoot tips.The shoot tips were transferred to solid culture medium（MS ＋ 0.5 mg · L^-1 Zeatin ＋ 0.1 mg · L^-1 NAA ＋ 1.0 mg · L^-1 GA3）for recovery and regeneration.The average survival rate and regeneration rate were 79.91% and 62.52%,respectively.There was no genetic variation in the regenerated plants based on assessment of SSR markers.
出处
《园艺学报》
CAS
CSCD
北大核心
2010年第9期1431-1438,共8页
Acta Horticulturae Sinica
基金
国家科技支撑计划项目(2006BAD13B10)
中央级公益性科研院所基本科研业务费专项(092060302-07)
农业部农作物种质资源保护利用专项
关键词
马铃薯
离体茎尖
小滴玻璃化法
超低温保存
遗传稳定性
potato
In vitro shoot tips
droplet vitrification method
cryopreservation
genetic stability
