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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 張孟基(Men-Chi Chang) | |
| dc.contributor.author | Row-Yu Ma | en |
| dc.contributor.author | 馬若瑀 | zh_TW |
| dc.date.accessioned | 2021-05-15T17:51:45Z | - |
| dc.date.available | 2016-08-25 | |
| dc.date.available | 2021-05-15T17:51:45Z | - |
| dc.date.copyright | 2014-08-25 | |
| dc.date.issued | 2014 | |
| dc.date.submitted | 2014-08-14 | |
| dc.identifier.citation | Abdrakhamanova A., Wang Q.Y., Khokhlova L., Nick P. (2003) Is microtubule disassembly a trigger for cold acclimation? Plant Cell Physiol., 44: 676–686
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/5087 | - |
| dc.description.abstract | 肌動蛋白去聚合因子 (Actin Depolymerizing Factor, ADF) 為一與肌動蛋白結合之蛋白質，其功能為藉由與actin結合將F-actin解聚為G-actin，在細胞中參與了細胞骨架之動態性變化。目前已知ADFs和植物細胞之生長發育、形態、細胞分裂、訊息傳導及生物性逆境相關，然而ADFs與植物非生物逆境耐受性之關連及其功能仍有待探討。因此本論文之實驗以異位表達之方式，利用35S啟動子過量表現水稻OsADF2及OsADF11基因於阿拉伯芥中，並觀察轉殖株與野生型外表型及其在非生物逆境下的反應。結果顯示T3同型結合子之OsADF2和OsADF11轉殖株在一般生長情況下轉殖株葉片大小較野生型阿拉伯芥為大，其抽苔時間與開花時間也較野生型為早。一般環境下發芽時間與野生型無差異。然而在150 mM NaCl的環境下，35S::OsADF2及35S::OsADF11轉殖株的發芽時間較野生型阿拉伯芥為早。另外將5天大及10天大之阿拉伯芥移植至含有150 mM NaCl之培養基後發現OsADF2及OsADF11轉殖株之根部生長及存活率皆大於野生型阿拉伯芥。另外以乾旱、高鹽、低溫逆境處理21天大之OsADF2及OsADF11轉殖株成株，結果顯示OsADF2及OsADF11皆較野生型耐旱，然而OsADF11在低溫處理後植物組織有受傷的情形發生，但此現象只在子葉部分觀察到，而新生葉則與野生型無明顯差異。利用紅外線熱影像檢測植株葉溫之變化，在一般條件下轉殖株與野生型葉溫無差異。另一方面以200 mM NaCl 處理三周大之OsADF2及OsADF11轉殖株及野生型阿拉伯芥，則發現OsADF2轉殖株之葉片溫度與野生型無顯著差異，而OsADF11轉殖株之葉溫則是較野生型為高。利用玉米ubiquitin promoter表現水稻OsADFs基因家族之GFP融合蛋白，接著以暫時性表達分析水稻OsADFs之次細胞定位，結果顯示水稻OsADF2、OsADF3及OsADF11與GFP之融合蛋白於細胞核及細胞質中皆有表現，其中OsADF2與GFP之融合蛋白在葉綠體中也有表現，而OsADF1、OsADF4、OsADF5、OsADF6與GFP之融合蛋白則是皆表現於葉綠體中。由以上之結果推論水稻OsADF2及OsADF11可能與植物面臨高鹽逆境以及乾旱逆境時的耐受機制有關。 | zh_TW |
| dc.description.abstract | Actin Depolymerizing Factors (ADFs) are small actin-binding proteins in cytoskeleton remodeling in cells. Recently, ADFs were reported to play a role in plant growth, development, cell division, signal transduction and pathogen resistance. However the functions of rice ADFs (OsADFs) under abiotic stress still remained to explore. In this thesis, we took heterologous gene expression approach to overexpress rice OsADF2 and OsADF11 in Arabidopsis, and observed the phenotype difference between transgenic and wild type plants under normal growth and abiotic stress. The phenotype of OsADF2 and OsADF11 transgenic plants showed bigger leaves size than Col-0, moreover, the bolting time and flowering time was earlier. The germination rate of transgenic plants had no difference compared to wild-type under normal condition while the germination rate of transgenic plants was faster after treatment with 150 mM NaCl. We also transferred the 5-day old and 10-day old seedlings to the 150 mM NaCl medium and found that the root growth and survival ratio of transgenic plants were both higher than wild-type. Next we grown 21-day old OsADF2, OsADF11 transgenic and wild-type plants under drought, salt and cold stresses, and OsADF2 and OsADF11 transgenic plants tended to be drought tolerance while injury tissue occurred in OsADF11 transgenic plant under cold treatment. Using non-destructive infrared thermography camera recorded the leaf temperature change under salt stress condition, the result showed that no obvious differences were found between transgenic and wild-type plants under normal condition. However OsADF11 transgenic plants showed higher leaf temperature compared to wild-type but not OsADF2 after treatment with 200 mM NaCl. In addition, to understand the subcellular localization of rice OsADFs, we used maize ubiquitin promoter to express rice OsADFs-GFP fusion protein through rice protoplast mediated transformation. The result showed that OsADF3-GFP and OsADF11-GFP were located in the nucleus and cytosol, and OsADF2-GFP was located in thel nucleus, the cytosol and also in chloroplasts. OsADF1, OsADF4, OsADF5 and OsADF6 were located in the chloroplasts. Taken together from above results, our study implied that rice OsADF2 and OsADF11 maybe involve in the plant response to drought and salt stress. | en |
| dc.description.provenance | Made available in DSpace on 2021-05-15T17:51:45Z (GMT). No. of bitstreams: 1 ntu-103-R01621103-1.pdf: 11733743 bytes, checksum: ac826c9564a612433563c388634eedd8 (MD5) Previous issue date: 2014 | en |
| dc.description.tableofcontents | 致謝 ii
中文摘要 iii Abstract v 表目錄 xi 圖目錄 xi 附錄目錄 xii 縮寫字對照 xiii 第一章 前言 1 第二章 前人研究 3 一、 植物在非生物逆境下之反應 3 二、 細胞骨架與肌動蛋白於植物之生長發育及非生物逆境耐受性 4 1. 植物細胞骨架之動態平衡 5 2. 細胞骨架之肌動蛋白感應外界環境因子及內生訊號 6 2. 1. 溫度 6 2. 2. 鹽分 8 2. 3. 乾旱 9 2. 4. 重金屬 10 三、 肌動蛋白去聚合因子(ADF) 10 1. 肌動蛋白去聚合因子 (ADF) 參與植物之生長發育 11 2. 肌動蛋白去聚合因子(ADF) 參與植物之生物逆境耐受性 12 3. 水稻肌動蛋白去聚合因子之相關研究 12 四、 研究目的及實驗架構 13 第三章 材料方法 15 一、 OsADF 基因家族之生物資訊分析 15 二、 試驗材料 15 1. 阿拉伯芥種子 15 2. 水稻種子 15 三、 阿拉伯芥生長條件及不同非生物逆境處理 16 1. 阿拉伯芥生長條件 16 2. 阿拉伯芥生理及外表型觀察之非生物性逆境處理條件 16 2.1 發芽率分析之逆境處理條件 16 2.2 根長分析及存活率之逆境處理條件 17 2.3 成株分析之逆境處理條件 17 2.4 遠紅外線熱感應影像分析鹽逆境處理轉殖株條件 17 四、 阿拉伯芥突變株及轉殖株之分子鑑定 18 1. 植物DNA之萃取 18 2. DNA洋菜瓊脂膠體電泳分析 19 五、 阿拉伯芥突變株及轉殖株之基因表現分析 19 1. RNA之抽取及製備 19 2. RNase Free DNase I 處理 19 3. cDNA 合成之反轉錄反應 20 4. 反轉錄聚合酶連鎖反應 (RT-PCR) 20 六、 阿拉伯芥轉殖株之質粒構築及逆境耐受性測試 20 1. 35S::OsADF2及35S::OsADF11 質粒構築流程 20 2. 異位表現(heterologous expression)水稻基因之阿拉伯芥轉殖 21 3. 大腸桿菌轉型作用 22 3.1 大腸桿菌勝任細胞製備 22 3.2 質粒大腸桿菌轉型 22 3.3 檢測轉型後大腸桿菌質粒colony PCR 22 3.4 大腸桿菌質粒DNA小量純化 23 4. DNA 片段的製備與回收 23 5. 阿拉伯芥基因轉殖 24 5.1 植物材料預備 24 5.2 農桿菌轉型 24 5. 2. 1 農桿菌勝任細胞製備 24 5. 2. 2 質粒農桿菌轉型 24 5. 2. 3 檢測轉型後之農桿菌質粒colony PCR 25 5. 2. 4 阿拉伯芥轉殖 25 6. 阿拉伯芥轉殖株篩選 26 6.1 抗生素篩選轉殖株 26 七、 OsADF基因家族之GFP融合蛋白之次細胞位置分析 26 1. Ub::OsADFs-GFP質粒構築流程 26 2. 水稻材料預備 27 3. 抽取水稻原生質體 27 4. 聚乙二醇(PEG) 轉殖水稻原生質體 27 5. 共軛焦顯微鏡 (confocal microscope) 觀察螢光蛋白表現 28 第四章 結果 29 一、 OsADF2及OsADF11基因於非生物逆境下之基因功能分析 29 1. 異位表現水稻OsADF2及OsADF11基因之阿拉伯芥轉殖株分子鑑定 29 2. 異位表現水稻OsADF2及OsADF11基因之阿拉伯芥轉殖株外表型觀察 29 2.1. 水稻OsADF2及OsADF11基因影響阿拉伯芥植株大小 29 2.2. 水稻OsADF2及OsADF11基因提早植株抽苔開花時間 30 3. 水稻OsADF2基因之基因功能性分析 30 3.1. 阿拉伯芥35S::OsADF2轉殖株之發芽率測定 30 3.2. 阿拉伯芥35S::OsADF2轉殖株於非生物逆境下之觀察 30 3.2.1. 發芽率分析 30 3.2.2. 根長分析 31 3.2.3. 成株觀察 31 4. 水稻OsADF11基因之基因功能性分析 33 4.1. 阿拉伯芥35S::OsADF11轉殖株之發芽率 33 4.2. 阿拉伯芥35S::OsADF11轉殖株於非生物逆境下之觀察 33 4.2.1 發芽率分析 33 4.2.2 根長分析 34 4.2.3 成株觀察 34 二、 水稻肌動蛋白去聚合因子家族 (Actin depolymerizing factor) 之GFP融合蛋白於水稻原生質體之次細胞定位 37 第五章 討論 38 一、 探討水稻OsADF2及OsADF11基因與植物生長發育之關係 38 1. 水稻OsADF2及OsADF11基因可能參與植物之生長發育及開花 38 二、 探討水稻OsADF2及OsADF11基因於非生物下之功能比較 40 1. OsADF2及OsADF11基因參與植物面臨高鹽逆境時的耐受性探討 42 2. OsADF2及OsADF11基因在低溫逆境下之反應探討 44 3. OsADF2及OsADF11基因參與植物面臨乾旱逆境時的耐受性探討 45 三、 水稻OsADF基因家族之GFP融合蛋白次細胞定位結果探討 47 四、 結語 48 參考文獻 49 表目錄 表一、 水稻肌動蛋白去聚合因子(OsADFs) 家族成員基因序號相關資訊 58 表二、 水稻肌動蛋白去聚合因子(OsADFs) 家族成員之次細胞定位 58 圖目錄 圖一、 阿拉伯芥過度表現水稻OsADF2及OsADF11轉殖株之分子鑑定與外表型分析 59 圖二、 阿拉伯芥過度表現水稻OsADF2及OsADF11轉殖株之抽苔及開花時間與野生型Col-0之分析比較 61 圖三、 阿拉伯芥過度表現水稻OsADF2於逆境下根長分析與存活率分析 63 圖四、 阿拉伯芥過度表現水稻OsADF11之於逆境下根長分析與存活率分析 65 圖五、 35S::OsADF2轉殖株成株之乾旱逆境觀察 67 圖六、 35S::OsADF2轉殖株成株之高鹽逆境觀察 69 圖七、 35S::OsADF2轉殖株成株之低溫逆境觀察 70 圖八、 35S::OsADF11轉殖株成株之乾旱逆境觀察 71 圖九、 35S::OsADF11轉殖株成株高鹽之逆境觀察 73 圖十、 35S::OsADF11轉殖株成株之低溫逆境觀察 74 圖十一、 紅外線熱感應影像分析阿拉伯芥過度表現水稻OsADF2及OsADF11轉殖株於200 mM NaCl處理之鹽份逆境下反應 75 圖十二、 水稻OsADFs 基因家族之GFP融合蛋白於水稻原生質體之次細胞定位 77 附錄目錄 附錄一、 水稻和阿拉伯芥之肌動蛋白去聚合因子家族胺基酸的序列比對及親緣演化樹分析 79 附錄二、 水稻肌動蛋白去聚合因子家族基因於不同組織與生長期之表現圖譜 80 附錄三、 基因轉殖表現載體構築之質粒圖 81 附錄四、 訊息傳遞分子與植物肌動蛋白骨架之關聯圖 82 附錄五、 本研究所使用之引子對一覽 83 附錄六、 與肌動蛋白相關之研究整理 84 | |
| dc.language.iso | zh-TW | |
| dc.subject | 非生物逆境 | zh_TW |
| dc.subject | 水稻肌動蛋白去聚合因子 | zh_TW |
| dc.subject | 細胞骨架 | zh_TW |
| dc.subject | 異位表達 | zh_TW |
| dc.subject | 植物生長發育 | zh_TW |
| dc.subject | abiotic stresses | en |
| dc.subject | Actin depolymerizing factor | en |
| dc.subject | Cytoskeleton | en |
| dc.subject | Heterologous expression | en |
| dc.subject | development and growth | en |
| dc.title | 水稻肌動蛋白去聚合因子OsADF2及OsADF11基因異位表達於阿拉伯芥中之功能性分析 | zh_TW |
| dc.title | Functional Analysis of Rice Actin Depolymerizing Factor OsADF2 and OsADF11 by Heterologous Expression in Arabidopsis | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 102-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 謝旭亮,趙光裕,洪傳揚,黃文理 | |
| dc.subject.keyword | 水稻肌動蛋白去聚合因子,細胞骨架,異位表達,植物生長發育,非生物逆境, | zh_TW |
| dc.subject.keyword | Actin depolymerizing factor,Cytoskeleton,Heterologous expression,development and growth,abiotic stresses, | en |
| dc.relation.page | 84 | |
| dc.rights.note | 同意授權(全球公開) | |
| dc.date.accepted | 2014-08-15 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 農藝學研究所 | zh_TW |
| 顯示於系所單位： | 農藝學系 | |
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