海南安诺兰菌根表面消毒方法研究

Study on Surface Disinfection of Mycorrhiza from Anota hannanensis in Hainan Province

采用75%乙醇、次氯酸钠溶液浸泡和无菌水洗涤的方法,对海南安诺兰的菌根进行表面消毒,以消毒强度逐步递减的方式,确定最佳分离植物内生真菌的表面消毒强度;同时设置漂洗液无菌检测和组织印迹无菌检测2种对照处理,确保适度的表面消毒强度,然后采用不同浓度的培养基分离培养内生真菌。结果表明,先用75%乙醇浸泡材料30 s,再用3.0%次氯酸钠浓度浸泡5 min处理,是海南安诺兰菌根内生真菌最佳的表面消毒方法,使用20%PDA双抗培养基、于25℃恒温条件下培养,能最大限度地分离所有植物内生真菌,分离效果最佳。

The surface disinfection method of mycorrhiza from Anota hannanensis were investigated. Plant materials were sterilized by surface treatment with 75% ethanol, sodium hypochlorite （NaClO）solution and sterilized water. The optimal surface sterilization intensity for isolating endophytic fungi was determined in a progressive decrease way with biological safety cabinet sterilization detection control,washing liquid sterilization detection and tissue blotting sterilization detection was controlled. The results showed that ,the best method of surface disinfection was with 75% ethanol teatment 30 s,then treating 15 rain with 3.0% NaClO solution.20% PDA medium supplemented with potassium benzylpenicillin and streptomycin sulfate at 25 ℃could maximum separat endophytic fungi and was more suitable for separation of endophytic fungi.