摘要
研究改进了Vierheilig等描述的AM菌根染色法:将根样于20%KOH溶液中60℃水浴透明40-120 min,5%醋酸酸化5min后,用5%醋酸墨水染色液(派克纯黑书写墨水Quink),于60℃水浴染色30 min,清水浸泡脱色(14h)后即可镜检。根皮层细胞内AM真菌的丛枝结构清晰可见,并且能够明确地分辨AM真菌与其它未知真菌。此外,Quink初染后,再经过SudanⅣ复染(60℃、60 min),70%乙醇脱色5min,暗隔真菌的透明菌丝内所积聚的脂类颗粒被SudanⅣ染上鲜红色,在复式显微镜下能够观察到此类透明菌丝在根皮层组织内的存在状况。采用甘油明胶为封固剂制片,根的染色效果可以保存长久。此项技术可以对同一种植物的多个根样进行同步的透明和染色处理,而且操作简便、低毒性、成本低廉、染色效果极佳,适用于野生和栽培草本植物AM菌根的染色和制片观察。
The procedure for staining arbuscular mycorrhizal（AM） fungal colonization in root tissues developed by Vierheilig et al.was modified by authors.Roots were cleared in 20% KOH for 40 to 120 min at 60℃ in water bath and then acidified by 5% acetic acid for 5 min.Cleared roots were stained for 30 min in 5% ink-vinegar solutions（Parker black writing ink,Quink） at 60℃ in water bath.Roots were destained by immersing in tap water for 14 h.Arbuscules of the AM fungi in root cortex were clearly visible and the distinctions between AM and other unidentified fungi could be noticed under a compound microscope.Moreover,lipid bodies within the hyaline hyphae of dark septate endophytes（DSE） were stained bright red and clearly distinguished when roots were restained in Sudan Ⅳ（60℃ for 60 min） and destained for 5 min in 70% ethanol.Stained roots could be mounted in glycerin jelly for making a permanent slide with high staining quality.This method should allow a large number of root samples of individual plant to be cleared and stained simultaneously and provided a simple,low toxic and inexpensive technique for staining AM fungi in cleared root for both wild and cultivated herbaceous plants with excellent staining results.
出处
《生态学报》
CAS
CSCD
北大核心
2010年第3期774-779,共6页
Acta Ecologica Sinica
基金
国家重点基础研究发展规划资助项目(2007CB106801)
国家自然科学基金资助项目(30571318,30600427)
关键词
丛枝菌根
醋酸墨水
暗隔真菌
arbuscular mycorrhiza ink-vinegar dark septate endophytes
