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摘要：植物内生真菌普遍认为是从表面已经灭菌的健康植物中分离纯化得到的，存在于健康植物的组织器官中，对植物本身不产生明显侵染的一类真菌[1]。随着研究的深入,人们发现植物内生真菌的次生代谢产物中含有各种不同的生物活性物质。目前己有多种植物内生真菌被研究出具有抑菌活性。本实验通过对采集的三十种不同的植物进行内生真菌的分离纯化，再通过真菌抑制实验筛选出具有抗青霉活性的内生真菌，然后提取目标内生真菌的DNA,PCR扩增其ITS并测序，并结合菌丝形态和产孢结构鉴定目标内生真菌的分类地位。通过该实验分离出了三种具有抑制真菌活性的内生真菌，分别是从桂花中分离得到的拟茎点霉Phomopsis sp.和曲霉属Aspergillus，从黄杨中分离到的水葫芦黑斑病菌Alternaria alternata。34671
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毕业论文关键词：内生真菌、分离、抑菌活性
Separation Purification and Identification of Antifungal Endophytic fungi in Plant
Abstract：Endophytic fungus is generally considered to be isolated and purified from healthy plants which the surface have been sterilized. They exist in healthy tissues and organs of plants, and they do not produce a significant infestation. As studying deep mycorrhizal fungi in plant, we found secondary metabolites in endophytic fungi contain various biologically active substances. Currently, antibacterial activity substances had been found in a variety of endophytic fungi. In this study, endophytic fungi were isolated and purified from thirty different plants. Antifungal activity fungi were screened through fungi inhibition experiments. Then, DNA of these endophytic fungi was purified. ITS was amplified and sequenced. Their sequence was blasted in NCBI database. At last, the classification status of these endophytic fungi wereidentified by the result of sequence and mycelium morphology. Three antifungal activity endophytic fungi were purified and identified in this experiment. They are Phomopsis sp. and Aspergillus from Osmanthus and Alternaria alternata from Boxwood.
Key words: Endophytic fungi, Separation, Antifungal activity
目录
摘要 I
目录 1
引言 3
1 材料与方法 3
1.1 实验材料 3
1.1.1 样品采集 3
1.1.2 药品 3
1.1.3 培养基 3
1.1.4 菌种 4
1.1.5主要的仪器设备 4
1.2 实验方法 5
1.2.1 样品采集 5
1.2.2 清洗 5
1.2.3 取样 5
1.2.4 表面灭菌 5
1.2.5 平板接种 5
1.2.6 室温培养 6
1.2.7 菌种纯化 6
1.2.8 菌种保藏 7
1.2.9 菌种发酵 7
1.2.10过滤发酵液 7
1.2.11抗真菌试验 8
1.2.12菌丝形态鉴定 9
1.2.13菌种DNA的提取、扩增、鉴定 9
2 结果 11
2.1 11号菌-拟茎点霉Phomopsis sp. 11
2.1.1 11号菌青霉抑制梯度实验结果 11
2.1.2 11号菌菌丝及产孢结构拍照 12
2.1.3 11号菌菌种鉴定 13
2.2 164号菌-曲霉属Aspergillus 13
2.2.1 164号菌青霉抑制梯度实验结果 13
2.2.2 164号菌菌丝及产孢结构拍照 13
2.2.3 164号菌菌种鉴定 15
2.3 268号菌-水葫芦黑斑病菌Alternaria alternata 15