中国中药杂志

2014, v.39(02) 197-203

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镰刀菌诱导结香对白木香叶内生真菌分布和群落构成的影响研究
Effects of agarwood formation induced by Fusarium sp. A2 on distribution and community composition of endophytic fungi in leaves of Aquilaria sinensis

高晓霞;周伟平;王磊;章卫民;严寒静;
GAO Xiao-xia;ZHOU Wei-ping;WANG Lei;ZHANG Wei-min;YAN Han-jing;Guangdong Pharmaceutical University;State Key Laboratory of Applied Microbiology / Ministry-Guangdong Province Jointly Breeding Base,South China, Guangdong Open Laboratory of Applied Microbiology,Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application,Guangdong Institute of Microbiology;

摘要(Abstract):

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。
The aim of the paper is to observe the distribution of the endophytic fungi in leaves of Aquilaria sinensis by using permanent paraffin-cut section,optical microscope photography and histochemistry. Total DNA was extracted with modified CTAB method and rDNA ITS regions of plant and endophytic fungi were amplified with eukaryotic universal primers. The rDNA ITS amplicon was characterized by RFLP analysis,sequencing of rDNA ITS library and phylogenetic analyses using PAUP by maximum parsimony. Fusarium sp. A2 was used to induce the formation of resinous in A. sinensis trees. As a result,endophytic fungi mainly distributed in spongy and phloem in leaf. Endophytic fungi distributed in the phloem in agarwood-producing tree and had a relatively high abundance. Phoma sp. and Collectrotrichum sp. were the absolute advantage species in the leaf tissues of non-resinous and agarwood-producing tree,respectively. Collectrotrichum sp. was the only fungal species detected both in the two types of A. sinensis with different levels of abundance. The culture-independent molecular method can be used to identify fungal species directly and rapidly from the plant tissues. Endophytic fungal communities in non-resinous and agarwood-producing A. sinensis leaf tissues were quite different.

关键词(KeyWords): 白木香;沉香形成;人工造香;内生真菌;rDNA ITS
Aquilaria sinensis;agarwood formation;artificial induce of agarwood;endophytic fungi;rDNA ITS

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(81102418,31100496);; 广东省中国科学院全面战略合作项目(2011B090300078);; 广东省科技计划项目(2012A030100014)

作者(Author): 高晓霞;周伟平;王磊;章卫民;严寒静;
GAO Xiao-xia;ZHOU Wei-ping;WANG Lei;ZHANG Wei-min;YAN Han-jing;Guangdong Pharmaceutical University;State Key Laboratory of Applied Microbiology / Ministry-Guangdong Province Jointly Breeding Base,South China, Guangdong Open Laboratory of Applied Microbiology,Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application,Guangdong Institute of Microbiology;

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