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为了研究沉香形成前后白木香叶组织中内生真菌的分布和群落构成,采用石蜡永久制片法观察白木香叶组织显微结构,细胞化学法确定内生真菌的分布。通过改良的CTAB法对白木香进行总DNA提取,利用真核生物通用引物同时对植物和内生真菌rDNA ITS序列进行PCR扩增,构建内生真菌rDNA ITS文库,RFLP分析和序列测定,用PUAP软件进行系统发育分析,构建最大简约树。结果表明:镰刀菌诱导结香前后,白木香内生真菌菌丝主要分布于海绵组织和韧皮部,结香后韧皮部内生真菌菌丝相对丰度明显增大;茎点霉是未结香白木香叶内生真菌优势种群,刺盘孢菌是已结香白木香叶内生真菌优势种群;刺盘孢菌是未结香和已结香白木香唯一共有内生真菌,且丰度差异较大。不依赖分离培养的分子生物学方法可快速直观地用于植物组织内生真菌的鉴定。白木香叶内生真菌具有一定的遗传多样性,沉香形成前后白木香内生真菌群落构成差异较大。
In order to study the distribution of endophytic fungi and the composition of the endophyte before and after the formation of the alopecuroides, the microstructure of the white leaf was observed by the permanent paraffin method, and the distribution of endophytic fungi was determined by cytochemical method. The total DNA was extracted from the white wood by modified CTAB method. The rDNA ITS library of endophytic fungi was amplified by PCR using common primers of eukaryotes. The rDNA ITS library, RFLP analysis and sequencing of endophytic fungi were constructed. The PUAP The software conducts phylogenetic analysis and constructs the largest parsimonious tree. The results showed that the mycelia of endophytic fungi were mainly distributed in the sponge and phloem before and after the Fusarium spp., And the relative abundance of endophytic fungi in the phloem was significantly increased after the fruiting; The dominant species of leaf endophytic fungi, Cercospora spp. Were the dominant species of endophytic fungi that had already ceased to be fragrant. The spore-forming bacteria were the only common endophytic fungus with un-jossed and fruiting bodies, and their abundance differed greatly . Molecular biology methods that do not rely on isolation and culture can be quickly and intuitively applied to the identification of endophytic fungi in plant tissues. Endophytic fungi of white wood have certain genetic diversity, and the composition of endophytic fungi community of white wood is different before and after the formation of agarwood.