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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 杜宜殷(Yi-Yin Do) | |
dc.contributor.author | Cheng-Ku Lin | en |
dc.contributor.author | 林政谷 | zh_TW |
dc.date.accessioned | 2021-06-08T05:07:09Z | - |
dc.date.copyright | 2011-07-18 | |
dc.date.issued | 2011 | |
dc.date.submitted | 2011-06-23 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/23682 | - |
dc.description.abstract | 苦瓜 (Momordica charantia L.) 是臺灣夏季主要蔬菜作物之一,為瞭解植物生長素及乙烯對苦瓜果實發育及後熟之影響。本論文針對苦瓜植物生長素受體基因McTIR1 (TRANSPORT INHIBITOR RESPONSE 1)、乙烯受體基因McETR1 (ETHYLENE RECEPTOR 1)及McERS1 (ETHYLENE RESPONSE SENSOR 1) 進行啟動子活性分析,於菸草及阿拉伯芥中觀察上述啟動子於各發育階段、逆境及生長調節劑誘導的基因表現情形。McTIR1啟動子於阿拉伯芥及菸草轉殖株之花萼、雄蕊和雌蕊等組織皆有表現;於菸草種皮、下胚軸、葉、莖頂、根尖及花瓣等其他部位亦可偵測到基因之表現。經創傷及黑暗培養處理,結果顯示啟動子活性可於轉殖株葉片及莖部受到誘導促進表現,而4°C低溫及乾旱逆境則會部分抑制啟動子活性於葉片之表現。處理ABA及不同種類植物生長素後,可部分抑制葉片中McTIR1啟動子活性,其中以ABA抑制效果較為顯著。另一方面,經分析基因組選殖系,取得苦瓜乙烯受體基因McETR1序列9,213 bp及McERS1序列7,313 bp,分析兩基因基因結構皆由7個顯子及6個隱子組成,啟動子序列分析結果顯示,兩啟動子皆含ABRE (ABA responsive element)、GARE (GA responsive element)、HSE (Heat stress responsiveness element)、CGCTA (MeJA- responsiveness) 及光反應序列等調控因子。McETR1啟動子於菸草轉殖株之分子驗證已完成,待取得菸草之轉殖純系種子。McETR1啟動子活性於阿拉伯芥子葉、莖部節間、側根、根尖、雄蕊、雌蕊及果莢表現。經不同植物生長調節劑處理阿拉伯芥轉殖株,ACC、IAA、GA和SA可誘導苦瓜乙烯受體基因McETR1啟動子於阿拉伯芥主根、側根及莖段連結節間處表現,以ACC處理誘導效果最佳。 | zh_TW |
dc.description.abstract | The bitter gourd (Momordica charantia L.) is one of prominent summer vegetable crop in Taiwan. In order to understand influence of auxin and ethylene on fruit development and ripening for bitter gourd, auxin receptor McTIR1 (TRANSPORT INHIBITOR RESPONSE 1) and ethylene receptor genes McETR1 (ETHYLENE RECEPTOR 1) and McERS1 (ETHYLENE RESPONSE SENSOR 1) from bitter gourd were analyzed for promoter activity in tobacco and Arabidopsis at different developmental stages and treatments with abiotic stresses and plant growth regulators. Promoter activity of McTIR1 was expressed in calyx, pistil and stamen of McTIR1pro::GUS transgenic Arabidopsis and tobacco, which also expressed in seed coat, hypocotyls, leaf, stem, shoot, root tip and petal of transgenic tobacco. The promoter activity was enhanced in leaf and stem of transgenic tobacco seedlings by wounding or under dark, but suppressed in leaf by 4oC and drought. Both ABA and auxins play the inhibitory role on McTIR1 promoter activity in leaf of transgenic tobacco seedlings, while the manifest inhibitory effect was contributed by ABA. On the other hand, 9,213 bp of McETR1 and 7,313 bp of McERS1 were obtained from genomic clones for ethylene receptor gene structure and promoter sequence. Both ethylene receptor genes from bitter gourd possess seven exons and six introns and several cis-acting elements, such as ABRE (ABA responsive element)、GARE (GA responsive element)、HSE (Heat stress responsiveness element) and CGCTA (MeJA-responsiveness). Gene integrity and copy number for McETR1::GUS transgenic tobacco has been analyzed by Southern hybridization. McETR1 promoter was expressed in cotyledon, stem internode, later root, root tip, pistil, stamen and siliques of transgenic Arabidopsis. When treated by ACC, IAA, GA, or SA, the promoter activity in McETR1::GUS transgenic seedling was enhanced in main root, later root, and stem internode with most obvious effect by ACC. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T05:07:09Z (GMT). No. of bitstreams: 1 ntu-100-R97628136-1.pdf: 3115874 bytes, checksum: 5512d1fc4790dd789a51b6942d62c572 (MD5) Previous issue date: 2011 | en |
dc.description.tableofcontents | 誌謝 ..............................................................................................I
中文摘要 ..........................................................................................II Abstract .............................................................................................III 壹、前言 .............................................................................................1 貳、前人研究 ......................................................................................2 一、植物生長素 ..........................................................................2 (一) 植物生長素調控基因傳導模式 .......................................2 (二) 泛素複合物SCFTIR1/AFB之調控 ………………...............3 (三) 植物生長素受體基因TIR1 .…………..…….………….4 (四) 植物生長素受體TIR1之基因表現 ..................................4 (五) 植物生長素受體AFBs (Auxin-Signaling F-box Proteins)..5 (六) 苦瓜植物生長素受體McTIR1之基因特性 ......................6 二、乙烯訊息傳導 .......................................................................7 (一) 乙烯受體 .......................................................................................7 (二) 乙烯受體蛋白特性 ......................................................................8 (三) 乙烯受體蛋白調控模式 ...............................................................9 (四) 阿拉伯芥及不同物種乙烯受體之基因表現 …............................10 (五) 苦瓜乙烯受體McETR1及McERS1之基因特性 .......................12 参、材料與方法 .........................................................................13 一、試驗材料 ...........................................................................13 (一) 苦瓜基因組庫及苦瓜乙烯受體cDNA ............................13 (二) 植物材料 .......................................................................13 (三) 試驗菌種材料 ...............................................................14 (四) 探針來源 .......................................................................14 (五) 基因結構分析之引子序列 ............................................14 (六) 轉殖之質體 ...................................................................15 二、試驗方法 ............................................................................15 (一) 南方氏雜交分析 ……………………………....................15 1. 噬菌體選殖系之限制酶圖譜分析 ....................................15 2. 基因組南方氏雜交分析 ……………………..…..……….17 (二) 次選殖 ..........................................................18 1. 大腸農桿菌勝任細胞製備 ……………….………………18 2. 膠體內DNA之回收與接合反應 ..............................18 3. 質體DNA之轉型 ..........................................18 4. 質體DNA小量製備 ........................................18 5. DNA 定序 .......................................................20 (三) 基因結構及啟動子序列分析 .......................................20 (四) 啟動子序列表現質體構築 ……………………………….20 1. pGKU/McETR1pro構築策略 ......................................20 2. pGKU/McERS1pro構築策略 .......................................23 (五) 聚合酶連鎖反應 ...........................................................26 (六) 質體大量製備 ...............................................................26 (七) 質體DNA純化 ..............................................................26 (八) 農桿菌電穿孔轉形 ………….........................................27 1. 農桿菌勝任細胞製備 ..................................................27 2. 農桿菌轉形反應 .........................................................28 3. 農桿菌質體 DNA之小量製備 ....................................28 (九) 基因槍暫時性表達 ........................................................29 1. 金屬粒子的製備 ………………………………..…..…….29 2. 基因槍暫時性表達 …………………………….……..…..30 (十) 阿拉伯芥之穩定性基因轉殖及篩選 …………………..….30 1. 農桿菌製備 .................................................................30 2. 阿拉伯芥之農桿菌轉殖 ............................................30 3. 阿拉伯芥轉殖株之抗生素篩選 ...................................31 (十一) 菸草之穩定性基因轉殖及篩選 .....................................31 1. 菸草植株製備 .............................................................31 2. 菸草葉圓片轉殖 ........................................................32 3. 停止共培養及篩選轉殖株 .........................................32 (十二) 轉殖株之啟動子活性分析 ……………………….…….....33 1. 不同發育階段之啟動子活性分析 …………………........33 2. 不同生長調節劑之逆境誘導處理 ………………………..33 3. 非生物逆境誘導處理 ……………………………………..34 (十三) GUS活性組織染色法 …………………..………………..35 肆、結果 …………....................................................................36 一、苦瓜植物生長素受體基因McTIR1啟動子活性分析 ….......36 (一) McTIR1啟動子活性於阿拉伯芥之穩定性分析................... 36 (二) McTIR1啟動子活性於菸草之穩定性分析.....................…..40 1. McTIR1啟動子轉殖株之分子驗證 ……………..………40 2. 不同發育階段之McTIR1啟動子活性分析 ……….......40 3. 不同逆境及植物生長調節劑誘導之McTIR1啟動子活性分析 ...........................................................................46 二、苦瓜乙烯受體基因啟動子活性分析 ...................................51 (一) McETR1、McERS1基因次選殖及序列分析 ........................51 (二) McETR1pro構築質體轉殖至蝴蝶蘭之暫時性表達分..........67 (三) McETR1pro構築質體轉殖至阿拉伯芥之穩定性分析..........67 1. 不同發育階段之McETR1啟動子活性分析 .................67 2. 不同逆境及植物生長調節劑誘導之McETR1啟動子活性分析 ..........................................................................73 (四) McETR1啟動子於菸草之穩定性分析 .............................73 伍、討論 ....................................................................................79 一、苦瓜植物生長素受體基因McTIR1啟動子活性分析 .............79 二、苦瓜乙烯受體基因啟動子活性分析 ......................................81 (一) 苦瓜乙烯受體基因啟動子序列分析 .................................81 (二) McETR1啟動子於阿拉伯芥之穩定性分析 ........................82 陸、結語 ...........................................................................................85 參考文獻 .........................................................................................86 | |
dc.language.iso | zh-TW | |
dc.title | 苦瓜植物生長素及乙烯受體基因之啟動子活性分析 | zh_TW |
dc.title | Promoter activity analysis of auxin and ethylene receptor genes from bitter gourd (Momordica charantia L.) | en |
dc.type | Thesis | |
dc.date.schoolyear | 99-2 | |
dc.description.degree | 碩士 | |
dc.contributor.coadvisor | 黃鵬林(Pung-Ling Huang) | |
dc.contributor.oralexamcommittee | 黃銓珍(Chang-Jen Huang),洪傳揚(Chwan-Yang Hong) | |
dc.subject.keyword | 苦瓜,植物生長素,乙烯,受體基因,啟動子活性分析, | zh_TW |
dc.subject.keyword | bitter gourd,auxin,ethylene,receptor,promoter activity, | en |
dc.relation.page | 94 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2011-06-24 | |
dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
dc.contributor.author-dept | 園藝學研究所 | zh_TW |
顯示於系所單位: | 園藝暨景觀學系 |
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