博碩士論文 105821024 詳細資訊




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姓名 李振嘉(Chen-Chia Lee)  查詢紙本館藏   畢業系所 生命科學系
論文名稱 台灣沉香樹相關的內生菌
(Endophytic microorganisms associated with agarwood trees in Taiwan)
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摘要(中) 沉香為一種昂貴的木材,應用於薰香與香水產業,為 Aquilaria spp. 或 Gyrinops spp. (瑞香科)的樹木受到真菌感染並處於病理條件下形成的一種黑色油樹脂。為了探討台灣台東縣太麻里鄉之沉香相關的內生菌,我們以消毒過的生長錐與刀片分別採集活體木材(可能含有沉香木和健康的木材)以及樹齡20歲左右的 A. sinensis 之樹幹底部的死沉香木進行內生菌的分離。菌株根據菌落型態和分子鑑定來區別真菌和細菌物種。在2017年2月到2018年2月的四次採集中,總共分離了617個菌株,並鑑定出34種真菌物種與7種細菌物種。其中 Lasiodiplodia hormozganensis (34個菌株,占總菌株數的5.5%)在每次採集都有出現,大多分離於死沉香木(32株)。 Trichoderma lentiforme (236個菌株,占總菌株數的38.3%)出現於其中三次採集,大多分離於健康的木材(175株)。另外 Aspergillus fijiensis, Fusarium solani species 28 和 Phaeoacremonium rubrigenum 也在四次採集中出現了兩次。而在細菌方面,大多數分離於死沉香木(Paenibacillus lautus,41個菌株,占總菌株數的6.7%)。在感染實驗中, 以 T. lentiforme 和 L. hormozganensis 作為病原菌感染兩歲大的 A. sinensis 。本研究利用內生真菌以人工誘導的方式,使 A. sinensis 產生沉香同時奠定基礎並促進台灣沉香產業的發展。
摘要(英) Agarwood is the most expensive wood product and used in the incense and perfume industry. It is a dark resinous wood that forms in Aquilaria and Gyrinops trees (Thymelaeaceae family) under pathological conditions, mainly fungal infection. In order to isolate endophytic microorganisms associated with agarwood at Taimali Township, Taitung County, Taiwan, samples were aseptically taken with an increment borer from living wood (containing presumably fresh agarwood and healthy wood), standardized borecores as well as cut off non-standardized agarwood from exposed old agarwood from living stems of a ca. 20-years-old plantation of A. sinensis. Microbial strains were classified to morphospecies and identified to species with specific molecular genetic markers depending on the systematic groups of fungi or bacteria. A total of 617 strains comprising 34 fungal and 7 bacterial species were identified from 4 collection events between February 2017 and February 2018. Strains of Lasiodiplodia hormozganensis (34 strains, 5.5% of total strains) were isolated at every collection event, 2 from discolored wood, 0 from non-agarwood and 32 from dead agarwood. Strains of Trichoderma lentiforme (236 strains, 38.3% of total strains) were isolated at three collections, 175 associated with discolored wood, 29 from non-agarwood and 32 from dead agarwood. Aspergillus fijiensis, Fusarium solani species 28 and Phaeoacremonium rubrigenum were isolated at two collection events. Bacteria (Paenibacillus lautus, 41 strains, 6.7% of total strains) were almost only isolated from dead agarwood. T. lentiforme and L. hormozganensis species identified as candidate pathogens were used in inoculation experiments in two-year-old A. sinensis. The study lays the foundation for the development of endophytic fungi for artificial infection methods in A. sinensis to induce agarwood and promote the agarwood industry in Taiwan.
關鍵字(中) ★ 沉香 關鍵字(英) ★ Aquilaria sinensis
★ Agarwood
★ Endophytic fungi
論文目次 摘要 i
Abstract ii
誌謝 iv
Contents v
List of figures viii
List of tables x
List of abbreviations xi
1. Introduction 1
1-1. The application of agarwood 1
1-2. The relationship of Aquilaria and Agarwood 1
1-3. The demand and conservation of Aquilaria species 2
1-4. Endophytic microorganisms associated with agarwood trees 3
1-5. The identification of endophytes 4
1-6. Research aims 5
2. Materials and methods 6
2-1. Sample locality and collection 6
2-1-1. Operation of increment borer 7
2-2. Isolation of microorganisms 8
2-3. DNA isolation 8
2-3-1. Tissue dissociation 8
2-3-2. Lysis 9
2-3-3. DNA binding 9
2-3-4. Wash 10
2-3-5. DNA elution 10
2-4. Polymerase Chain Reaction (PCR) 11
2-4-1. Agarose gel electrophoresis 11
2-4-2. DNA sequencing 12
2-5. Identification of microorganisms 12
2-5-1. Morphological identification 12
2-5-2. Molecular identification 12
2-6. Identification of Trichoderma species 13
2-6-1. Morphological characterizations 13
2-6-2. Illustrations of micromorphological characteristics 14
2-6-3. Growth-rate trials 14
2-7. Media 15
2-8. Inoculation experiments 15
3. Results 17
3-1. Taxonomic treatment of important species 17
3-1-1. The identification of Aquilaria species 17
3-1-2. Trichoderma lentiforme 20
3-1-3. Aspergillus fijiensis 25
3-1-4. Phaeoacremonium rubrigenum 27
3-1-5. Lasiodiplodia hormozganensis 29
3-1-6. Fusarium solani species 28 33
3-2. Isolation of bacteria in association with Aquilaria sinensis 35
3-2-1. Paenibacillus lautus 36
3-3. Inoculation experiments 38
3-4. Species richness of endophytic microorganisms 39
4. Discussion 40
4-1. Comparison of standardized and non-standardized sampling 40
4-2. Bacteria in agarwood 41
4-3. Microorganisms from standardized sampling 41
4-4. Identification of microorganisms 43
4-5. Possible roles of the two most common microorganisms (Trichoderma lentiforme and Lasiodiplodia hormozganensis) 44
4-6. Inoculation experiment 46
5. References 47
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指導教授 羅南德 孫維欣(Roland Kirschner Wei-Hsin Sun) 審核日期 2019-1-17
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