[关键词]
[摘要]
目的为罗汉果Siraitia grosvenorii种质资源的保存提供一条新途径。方法以继代5d生长健壮的罗汉果试管苗0.8~cm长嫩梢进行预培养,剥取2~3mm茎尖,25℃下60%PVS2装载,再用00%PVS2在0℃脱水处理,更换新鲜00%PVS2后投入液氮保存24h,化冻,用MS+.2mol/L蔗糖的液体培养基洗涤,滤纸吸干后接种于MS+.0mg/L6-BA+0.05mg/L NAA+0.mg/L GA3+0.8%琼脂粉+45g/L蔗糖的恢复培养基上,25℃暗培养7d,转入正常光下培养。再生苗生根培养基为/2MS+0.2mg/L NAA+30g/L蔗糖。结果嫩梢于MS+0.7mol/L蔗糖的培养基上预培养3d,茎尖装载40min,脱水50min,液氮保存后于40℃水浴中快速化冻,洗涤40min,恢复培养周时存活率最高可达00%,30d时再生率最高可达78.33%。再生苗转入生根培养基可形成完整植株。结论罗汉果种质资源的玻璃化法超低温保存操作简单、成活率高、再生植株正常。
[Key word]
[Abstract]
Objective To get a new approach for conserving the germplasm of Siraitia grosvenorii Methods Shoots, 0.8-1 cm in length, excised from test-tube plantlets which were subcultured 15 d, were precultured. Then its shoot-tips about 2-3 mm in length were dissected and loaded with 60% PVS2 at 25 ℃, and dehydrated with 100% PVS2 at 0 ℃, changed for fresh 100% PVS2 prior to directly plunging into liquid nitrogen. After cryopreservation for 24 h, the shoot tips were thawed, rinsed in MS+1.2 mol/L sucrose medium, blotted with filter papers, then plated on the MS + 1.0 mg/L 6-BA+0.05 mg/L NAA + 0.1 mg/LGA3 + 0.8% agar + 45 g/L sucrose medium at 25 ℃ for 7 d in dark prior to exposure to the light. The root medium for regeneration plantlets was 1/2 MS + 0.2 mg/L NAA + 30 g/L sucrose. Results Shoots were precultured for 3 d on MS + 0.7 mol/L sucrose medium, then its shoot-tips loaded for 40 min, dehydrated for 50 min. After cryopreservation, the shoot tips were rapidly thawed in water at 40 ℃, then rinsed for 40 min. The survival rate of shoot-tips plated on the recovering medium in one week was 100%, and regeneration rate after 30 d was 78.33%, which was the highest. The regeneration plantlets inoculated on root medium were reconstructed integrating plantlets. Conclusion The method of vitrification to cryopreserve the germplasm of S. grosvenorii is a simple way with high survival rate and normal regeneration plants.
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[基金项目]
广西自然科学基金资助项目(桂科自0542032);广西师范大学博士启动基金项目;广西研究生教育创新计划项目(20080602070M255)