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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 曲芳華(Fang-Hua Chu) | |
| dc.contributor.author | Hui-Ling Hsieh | en |
| dc.contributor.author | 謝慧玲 | zh_TW |
| dc.date.accessioned | 2021-06-15T03:58:35Z | - |
| dc.date.available | 2010-05-20 | |
| dc.date.copyright | 2010-05-20 | |
| dc.date.issued | 2010 | |
| dc.date.submitted | 2010-05-17 | |
| dc.identifier.citation | 王升陽 (2006) 木材香味成分研究之新進展. 林業研究專訊 13(4): 5-9
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Plant J 27: 315-324 Lücker J, Bowen P, Bohlmann J (2004) Vitis vinifera terpenoid cyclases: functional identification of two sesquiterpene synthase cDNAs encoding (+)-valencene synthase and (-)-germacrene D synthase and expression of mono- and sesquiterpene synthases in grapevine flowers and berries. Phytochemistry 65: 2649-2659 Maia BH, de Paula JR, Sant’Ana J, da Silva MF, Fernandes JB, Vieira PC, Costa Mdo S, Ohashi OS and Silva JNM (2000) Essential oils of Toona and Cedrela species (Meliaceae): Taxonomic and ecological implications. J Braz Chem Soc 11: 629-639. Martin DM, Faldt J, Bohlmann J (2004) Functional characterization of nine Norway Spruce TPS genes and evolution of gymnosperm terpene synthases of the TPS-d subfamily. Plant Physiol 135: 1908-1927 Mumm R, Hilker M (2005) The significance of background odour for an egg parasitoid to detect plants with host eggs. 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Nat Prod Res 22: 360-364 Ribeiro VL, Rolim V, Bordignon S, Henriques AT, Dorneles GG, Limberger RP, von Poser G (2008) Chemical composition and larvicidal properties of the essential oils from Drimys brasiliensis Miers (Winteraceae) on the cattle tick Rhipicephalus (Boophilus) microplus and the brown dog tick Rhipicephalus sanguineus. Parasitol Res 102: 531-535 Runyon JB, Mescher MC, De Moraes CM (2006) Volatile chemical cues guide host location and host selection by parasitic plants. Science 313: 1964-1967 Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4: 406-425 Schmidt CO, Bouwmeester HJ, Bulow N, Konig WA (1999) Isolation, characterization, and mechanistic studies of (-)-alpha-gurjunene synthase from Solidago canadensis. Arch Biochem Biophys 364: 167-177 Starks CM, Back K, Chappell J, Noel JP (1997) Structural basis for cyclic terpene biosynthesis by tobacco 5-epi-aristolochene synthase. 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Plant Cell 12: 2237-2246 Wang SY, Wang YS, Tseng YH, Lin CT, Liu CP (2006) Analysis of fragrance compositions of precious coniferous woods grown in Taiwan. Holzforschung 60: 528-532 Wang SY, Wu CL, Chu FH, Chien SC, Kuo YH, Shyur LF, Chang ST (2005) Chemical composition and antifungal activity of essential oil isolated from Chamaecyparis formosensis Matsum. wood. Holzforschung 59: 295-299 Williams DC, McGarvey DJ, Katahira EJ, Croteau R (1998) Truncation of limonene synthase preprotein provides a fully active 'pseudomature' form of this monoterpene cyclase and reveals the function of the amino-terminal arginine pair. Biochemistry 37: 12213-12220 | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/44928 | - |
| dc.description.abstract | 臺灣杉 (Taiwania cryptomerioides Hayata) 為臺灣原生針葉樹種，近年來對其二次代謝萃取產物成分之生物活性有諸多研究；若能從分子生物學角度了解其生合成酵素，將使人為調控代謝產物成分的可能性增加。本研究鎖定萜類產物中之倍半萜，搜尋資料庫中針葉樹倍半萜類合成酶基因序列，設計簡併性引子對，利用反轉錄聚合酶連鎖反應技術，自成熟樹木之葉組織釣取到一具有典型萜類合成酶保守序列之基因，再經由5’及3’端快速增幅cDNA端點 (rapid amplification of cDNA ends, RACE) 選殖，共獲得基因全長1791 bp，轉譯後蛋白質分子量為70.2 kDa。此基因主要於幼葉、雌花及毬果中表現；另外，將葉部經水楊酸浸泡處理後，基因表現量有增加的現象。將此基因構築於蛋白質表現之載體中，轉入大腸桿菌 (Escherichia coli) 表現，發現此蛋白質可催化倍半萜之前驅物，生成倍半萜α-Gurjunene，故將此基因命名為Tc-Gur。為了解該基因在植物的表現情形，以基因轉殖方式表現，發現阿拉伯芥同樣會生成α-Gurjunene，而在臺灣杉毛狀根則生合成不同的倍半萜產物α-Muurolene。 | zh_TW |
| dc.description.abstract | Taiwania (Taiwania cryptomerioides Hayata) is a native conifer species of Taiwan. Many studies have demonstrated the bioactivities of secondary metabolites extracted from Taiwania in recent years. To regulate the biosynthsis of metabolites in Taiwania, it should be illustrate the synthesis based on the molecular level. In this study, we focus on sesquiterpene synthase of Taiwania. First, a pair of degenerated primers was designed for reverse transcription-polymerase chain reaction (RT-PCR) using total RNA extracted from leaves of mature tree. A DNA fragment with the conserved region of the terpene synthases (TPSs) was obtained. After 5’ and 3’ Rapid Amplification of cDNA Ends (RACE), the full-length cDNA was obtained, and it contains an open reading frame of 1791 bp with a predicated molecular mass of 70.2 kDa. The gene was highly expressed in young leaves, female flowers and cones. The expression in leave was enhanced after the treatment of soaking in 100 μM salicylic acid (SA). To identify the function of the obtained TPS, the recombinant protein from Escherichia coli was incubated with farnesyl diphosphate (FPP). After GC/MS analysis and retention time as well as mass spectrum matching with authentic standards revealed that the major product of TPS was sesquiterpene α-Gurjunene. The gene was named as Tc-Gur. To investigate the synthesis of this TPS in plant system, α-Gurjunene is also synthesized in transgenic Arabidopsis. But another sesquiterpene, α-Muurolene, is synthesized in transgenic hairy root of Taiwania. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-15T03:58:35Z (GMT). No. of bitstreams: 1 ntu-99-R96625035-1.pdf: 1202695 bytes, checksum: 1f15ce56a2a3f79b857bac5344f31ec2 (MD5) Previous issue date: 2010 | en |
| dc.description.tableofcontents | 口試委員審定書 I
誌謝 II 摘要 III Abstract IV 目錄 VI 圖目錄 IX 表目錄 X 一、前言 1 1.1. 臺灣杉 1 1.2. 萜類 2 1.3. 萜類合成酶序列特徵 4 二、材料方法 8 2.1. 植物材料與生長條件 8 2.2. 總量RNA之萃取 8 2.3. cDNA製備 8 2.4. 釣取基因 9 2.4.1. 設計簡併性引子對 9 2.4.2. 釣取基因片段 9 2.4.3. 基因全長之選殖 10 2.5. 序列比對與演化分析 11 2.6. 蛋白質結構預測 12 2.7. 蛋白質表現 12 2.7.1. 構築表現載體 12 2.7.2. 誘導表現重組蛋白質 13 2.7.3. 蛋白質電泳 14 2.7.4. 免疫雜合反應 14 2.8. 活性反應 15 2.8.1. 蛋白質之生成及反應 15 2.8.2. 氣相層析質譜分析儀 (GC/MS) 進行產物鑑定 16 2.9. 檢測在各組織mRNA表現量 17 2.9.1. 反轉錄聚合酶連鎖反應 (RT-PCR) 17 2.9.2. 即時定量PCR (real-time quantitative PCR) 17 2.10. 傷害處理 18 2.10.1. 化學性傷害處理 18 2.10.2. 物理性創傷處理 18 2.10.3. 反轉錄聚合酶連鎖反應 (RT-PCR) 18 2.10.4. 即時定量PCR (real-time quantitative PCR) 18 2.11. 阿拉伯芥之轉殖 19 2.11.1. 轉殖載體之構築 19 2.11.2. 轉殖株之篩選 20 2.11.3. GC/MS分析阿拉伯芥轉殖株之產物 20 2.12. 臺灣杉毛狀根之轉殖 21 2.12.1. GC/MS分析毛狀根轉殖體之產物 21 2.12.2. 毛狀根轉殖體產物之定量分析 21 三、實驗結果 22 3.1. 基因全長之選殖 22 3.2. 序列比對與演化分析 25 3.3. 蛋白質結構預測 29 3.4. 表現重組蛋白質 30 3.5. 重組蛋白質活性反應 31 3.6. Tc-Gur於各組織基因表現量 32 3.7. 經創傷、浸泡處理之Tc-Gur表現量 34 3.8. 阿拉伯芥之基因轉殖 36 3.9. 臺灣杉毛狀根之基因轉殖 37 四、討論 38 4.1. Tc-Gur基因序列特性 38 4.2. Tc-Gur生合成產物分析 39 4.3. Tc-Gur生合成產物可能之生物活性 42 4.4. 組織特異性 43 4.5. 化學性傷害與物理性創傷處理對基因表現的影響 44 五、結論 45 六、引用文獻 47 七、附錄 55 | |
| dc.language.iso | zh-TW | |
| dc.subject | 水楊酸 | zh_TW |
| dc.subject | 臺灣杉 | zh_TW |
| dc.subject | 倍半萜 | zh_TW |
| dc.subject | 合成酶 | zh_TW |
| dc.subject | α-Gurjunene | zh_TW |
| dc.subject | α-Muurolene | zh_TW |
| dc.subject | sesquiterpene synthase | en |
| dc.subject | Taiwania cryptomerioides Hayata | en |
| dc.subject | salicylic acid | en |
| dc.subject | α-Muurolene | en |
| dc.subject | α-Gurjunene | en |
| dc.title | 臺灣杉倍半萜合成酶基因之選殖與表現 | zh_TW |
| dc.title | Cloning and Expression of a Sesquiterpene Synthase Gene from Taiwania cryptomerioides Hayata | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 98-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 孟孟孝(Meng-Hsiao Meng),陳振榮(Zenn-Zong Chen),張淑華(Shu-Hwa Chang),王升陽(Sheng-Yang Wang) | |
| dc.subject.keyword | 臺灣杉,倍半萜,合成酶,α-Gurjunene,α-Muurolene,水楊酸, | zh_TW |
| dc.subject.keyword | Taiwania cryptomerioides Hayata,sesquiterpene synthase,α-Gurjunene,α-Muurolene,salicylic acid, | en |
| dc.relation.page | 55 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2010-05-18 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 森林環境暨資源學研究所 | zh_TW |
| 顯示於系所單位： | 森林環境暨資源學系 | |
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