摘要：植物内生菌是一个具有多样性的微生物类群，代谢产物特别的丰富，目前以抗菌、抗肿瘤等指标的药物开发为主要研究方向。本论文从植物中分离内生菌，进行了植物标本采集、内生菌分离纯化、抗菌活性植株筛选、活性菌株形态鉴定、菌种鉴定等研究，为进一步得到天然抗菌活性物质提供条件。本人从6种植物样本中分离纯化获得54株内生真菌、5种内生细菌。将内生真菌发酵处理液经过细菌筛选器筛选后，离心取上清，采用管碟法进行抑菌活性筛选；同样将内生细菌液体培养基培养液离心取上清，采用管碟法进行抑菌活性筛选。在本试验条件下，编号为HS-D2、LM-A4、LM-2的细菌培养液对青霉菌有很强的抑制作用，其抑菌圈直径分别为28mm、26mm、12mm。最后，对抑真菌效果较强的内生细菌LM-A4、HS-D2、LM-2菌株进行菌落特征培养、形态鉴定以及16S rDNA测序。综上所述，判定这三种内生细菌分别为甲基营养芽孢杆菌、解淀粉芽孢杆菌、内生多粘类芽孢杆菌。76472

毕业论文关键词：植物内生细菌；分离纯化；抗菌活性；鉴定

Isolation Purification and Identification for Inhibiting Fungi of the Endophytes 

Abstract: Endophyte is a special and important group of microorganisms with taxonomic persity, which can produce various bioactive substances。 At present, the ma -in research direction is the development of antibiotics, such as antibacterial, antitumor and other indicators。 The following items were carried through in this research: collection of plant samples, surface sterilization of plant samples, isolation and purification of endophytes, morphological identification of active, screening for antibacterial endophytes and identification of the active strain, which can be resources for natural antimicrobial products。 Their secondary metabolites were studied。 Fifty-four Endophytic Fungi strains and five Endophytic Bacteria strains were isolated from six sorts of plants。 Endophytic Fungi fermentation fluid was disposed by centrifugation。 Endophytic Bacteria fermentation fluid was disposed by centrifugation。 Supernatant liquid was used for antibacterial tests by Cylinder Plate Method。 The processed fermentation fluid of endophytic bacteria HS-D2、LM-A4 and LM-2 strain derived from Arachis hypogaea Linn and Chimonanthus praecox(Linn。) Link showed strong inhibition to bgbfngnvb。nhg, and the inhibition zone dimeter of the processed fermentation fluid of endophytic bacteria HS-D2、LM-A4 and LM-2 against bgbfngnvb。nhg were about 28mm、26mm、12mm。 Finally, antibacterial endophytes strain LM-A4、HS-D2and LM-2 were identified by cultivation characters, morphologic observations and sequence analysis of the 16SrDNA as the one of Bacillus Methylotrophicus、Bacillus Amyloliquefaciens、Paenibacillus Polymyxa。

Keywords: endophytic bacteria; isolation and purification; antibacterial activity; identifyication

目录

引言 1

1 材料与方法 2

1。1实验设备 2

1。2实验试剂和药品 2

1。2。1试剂 2

1。2。2药品 3

1。3培养基 3

1。3。1 PDA液体培养基 3

1。3。2 PDA固体培养基 3

1。3。3 LB液体培养基 3

1。3。4 LB固体培养基 4

1。4内生菌的分离 4

1。4。1植物样本的采集