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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林盈仲 | zh_TW |
| dc.contributor.advisor | Ying-Chung Jimmy Lin | en |
| dc.contributor.author | 劉香伶 | zh_TW |
| dc.contributor.author | Hsiang-Lin Liu | en |
| dc.date.accessioned | 2023-08-30T16:18:04Z | - |
| dc.date.available | 2023-11-10 | - |
| dc.date.copyright | 2023-08-30 | - |
| dc.date.issued | 2023 | - |
| dc.date.submitted | 2023-07-14 | - |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/89199 | - |
| dc.description.abstract | 木材是植物維管形成層增生分化出的次生木質部，由次級細胞壁所組成。次級細胞壁的主成分之一是木質素，木質素增加了細胞壁的剛性和疏水性，卻也造成木材在應用中生質轉化的困難。木質素由三種單木質醇所組成，目前單木質醇的生合成途徑已被揭示，透過研究單木質醇生合成的轉錄調控，可以在降低木質素含量的情況下，避免對植物的生長發育造成負面影響。在對轉錄調控網路的研究中人們知道轉錄因子除了直接與啟動子結合調控基因表達外，轉錄因子之間也會形成轉錄因子複合體去影響基因的轉錄。而儘管在阿拉伯芥 (Arabidopsis thaliana) 中對次級細胞壁生合成相關的研究有許多，但研究木本植物中木材生合成轉錄調控的部分卻很少，在過去的研究中是有找到在毛果楊 (Populus trichocarpa) 中與單木質醇生合成基因啟動子結合之轉錄因子。然而這只是初步讓我們知道這些轉錄因子與單木質醇生合成相關，但尚未清楚這些轉錄因子會如何形成複合體調節基因表達，進而影響木材形成。故本研究中以高通量減數分裂法酵母菌雙雜合系統 (meiosis-directed yeast two-hybrid) 篩選與單木質醇生合成基因啟動子結合之轉錄因子，與在木質部特異性表達之轉錄因子之間的交互作用。並以菌落計數分析儀來量化菌落大小，再透過GateMultiplex分析量化的結果。之後為確認交互作用在生物體中是否也會實際發生，通過原生質體中進行雙分子螢光互補 (bimolecular fluorescence complementation)，進一步驗證了篩選自m-Y2H的陽性交互作用在體內 (in vivo) 的表現。最終本研究篩選並驗證3個單木質醇生合成相關轉錄因子與8個木質部特異性表達轉錄因子之間17組交互作用。 | zh_TW |
| dc.description.abstract | Wood, also known as secondary xylem, is composed of secondary cell walls (SCWs). One of the main components of the secondary cell wall is lignin, and lignin increases rigidity and hydrophobicity of cell wall, but also causes the problem in wood applications. Lignin is composed of three types of monolignols, and the biosynthesis pathway of monolignol has been revealed. Through the study of transcriptional regulatory networks, transcription factors may form the transcription factor complexes to regulate gene expressions. While there are many studies related to SCW synthesis in Arabidopsis thaliana, there are few studies on transcriptional modulations of wood biosynthesis in woody plants. The previous study found that the transcription factors bind to the promoters of monolignol biosynthesis-related genes in Populus trichocarpa. However, it is not clear how these monolignol biosynthesis-related transcription factors form complexes to regulate gene expression in xylem. This study used high-throughput meiosis-directed yeast two-hybrid (m-Y2H) to detect the interactions between monolignol biosynthesis-related transcription factors and xylem-specific transcription factors. PhenoBooth was used to capture the images of the plates, and GateMultiplex software was used to analyze the quantified results. The positive interactive combinations from m-Y2H were further verified in vivo by bimolecular fluorescence complementation (BiFC) in protoplasts. We total identified 17 interactions between 3 monolignol biosynthesis-related transcription factors and 8 xylem-specific expressed transcription factors. | en |
| dc.description.provenance | Submitted by admin ntu (admin@lib.ntu.edu.tw) on 2023-08-30T16:18:04Z No. of bitstreams: 0 | en |
| dc.description.provenance | Made available in DSpace on 2023-08-30T16:18:04Z (GMT). No. of bitstreams: 0 | en |
| dc.description.tableofcontents | 口試委員審定書 i
誌謝 ii 中文摘要 iii Abstract iv 壹、前言 1 貳、材料與方法 6 (一) m-Y2H之實驗原理 6 (二) m-Y2H中baits和preys之選擇 6 (三) m-Y2H之BD文庫和AD文庫之建立 6 (四) 酵母菌轉殖 7 (五) 酵母菌菌落PCR 8 (六) 自體活化測試 9 (七) m-Y2H實驗流程 10 (八) 使用ROTOR HDA在m-Y2H進行高通量篩選 11 (九) BiFC構築體建立 13 (十) 毛果楊原生質體純化 14 (十一) 原生質體轉染 14 參、實驗結果 16 (一) TF-TF之間交互作用之篩選 16 (二) BiFC驗證篩選出的TF-TF組合在生物體內的交互作用 20 肆、討論 24 (一) m-Y2H篩選TFs之間交互作用結果 24 (二) TFs形成複合體後對基因的調控 25 (三) BiFC中EYFP訊號和mCherry訊號是否重疊對轉錄調控的意義 26 (四) 利用酶催化鄰近標記進一步驗證交互作用並進行後續蛋白質分析 27 (五) 透過同位素標記苯丙胺酸來觀察TF對單木質醇生合成路徑中化合物通量的影響 28 伍、參考文獻 30 陸、圖附錄 37 柒、表附錄 70 | - |
| dc.language.iso | zh_TW | - |
| dc.subject | 雙分子螢光互補 | zh_TW |
| dc.subject | 高通量 | zh_TW |
| dc.subject | 減數分裂法酵母菌雙雜合系統 | zh_TW |
| dc.subject | 轉錄因子複合體 | zh_TW |
| dc.subject | 單木質醇生合成 | zh_TW |
| dc.subject | 轉錄因子 | zh_TW |
| dc.subject | monolignol biosynthesis | en |
| dc.subject | meiosis-directed yeast two-hybrid | en |
| dc.subject | transcription factor complexes | en |
| dc.subject | High-throughput | en |
| dc.subject | BiFC | en |
| dc.subject | transcription factor | en |
| dc.title | 透過減數分裂法酵母菌雙雜合系統鑑定單木質醇生合成轉錄因子複合體 | zh_TW |
| dc.title | Identification of transcription factor complexes for monolignol biosynthesis by meiosis-directed yeast two-hybrid | en |
| dc.type | Thesis | - |
| dc.date.schoolyear | 111-2 | - |
| dc.description.degree | 碩士 | - |
| dc.contributor.oralexamcommittee | 張典顯;李金美;麥德倫 | zh_TW |
| dc.contributor.oralexamcommittee | Tien-Hsien Chang;Chin-Mei Lee;Te-Lun Mai | en |
| dc.subject.keyword | 高通量,減數分裂法酵母菌雙雜合系統,轉錄因子複合體,單木質醇生合成,轉錄因子,雙分子螢光互補, | zh_TW |
| dc.subject.keyword | High-throughput,meiosis-directed yeast two-hybrid,transcription factor complexes,monolignol biosynthesis,transcription factor,BiFC, | en |
| dc.relation.page | 74 | - |
| dc.identifier.doi | 10.6342/NTU202301384 | - |
| dc.rights.note | 同意授權(限校園內公開) | - |
| dc.date.accepted | 2023-07-17 | - |
| dc.contributor.author-college | 生命科學院 | - |
| dc.contributor.author-dept | 植物科學研究所 | - |
| dc.date.embargo-lift | 2028-07-06 | - |
| 顯示於系所單位： | 植物科學研究所 | |
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