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研究生:張芯
研究生(外文):Sim Chong
論文名稱:番茄ERF36與ERF39在逆境反應之功能研究
論文名稱(外文):Functional study of tomato ERF36 and ERF39 in stress responses
指導教授:鄭秋萍鄭秋萍引用關係
指導教授(外文):Chiu-Ping Cheng
口試委員:詹明才葉開溫鄭石通
口試委員(外文):Ming-Tsair ChanKai-Wun YehShih-Tong Jeng
口試日期:2013-07-30
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:植物科學研究所
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2013
畢業學年度:101
語文別:中文
論文頁數:76
中文關鍵詞:乙烯反應轉錄因子青枯病乾旱逆境鹽害逆境
外文關鍵詞:Ethylene-response factors (ERFs)bacteria wilt (BW)droughtsalt stress
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植物在自然環境中常遭受各種逆境脅迫,造成植物生長與農作物產量影響極巨,例如,由青枯病菌(Ralstonia solanacearum, Rs)引起的青枯病(bacterial wilt, BW)與缺水逆境都會導致植物急速死亡,造成農業上重大損失。植物為了生存演化出一套複雜的調控機制以調控並抵禦逆境反應,而乙烯轉錄因子(ethylene-response factor, ERFs)是植物特有的轉錄因子且參與許多逆境反應。我們先前的研究顯示在番茄中的兩個屬於VIII群ERF的成員SlERF36與SlERF39可能在青枯病與乾旱防禦反應具一定的重要性,故本研究旨在利用過量表現與基因靜默的轉殖植物研究策略進一步分析這兩個基因在植物逆境反應之確切功能與其調控機制。由結果發現SlERF36蛋白確實具有抑制轉錄活性的功能;過量表現SlERF36可能會使轉殖菸草對青枯病感病些微增加、對滲透壓逆境及乾旱抗性降低,推測SlERF36可能抑制抗病與抗旱相關啟動基因(activators)表現而造成這些性狀。另外,研究結果顯示SlERF39具有抑制轉錄活性的功能;過量表現SlERF39對青枯病抗性降低、對滲透壓及乾旱逆境反應降低,而較抗鹽害逆境,推論SlERF39可能使ET與JA訊息傳遞之抑制基因(repressors)表現降低進而增加ET與JA表現。SlERF39可能抑制促進基因表現進而對缺水逆境抗性降低,而藉由抑制啟動基因表現對鹽害逆境抗性增加。未來可利用本研究所得之過量表現與基因靜默之轉殖植物,更深入探討SlERF36與SlERF39在各種逆境反應之功能,希望有助於釐清對植物病害與非生物逆境之複雜的交互作用,並增加對這些逆境防禦機制的了解並加以運用。

Plant constantly encounters environmental stresses such as drought, salt, heat even pathogens and pest, which effect plant growth and crop productivity. Bacterial wilt (BW), which is caused by Ralstonia solanacearum, is a very complex deadly disease and shares certain common features with water stress responses induced plant death and crop loss. Hence, the plants have evoluted a complex mechanism to regulate and defense to stress responses. Ethylene response factors (ERFs) is a large family of plant-specific transcription factors involved in various stress responses. Our previous study revealed that two Group VIII ERFs from tomato, namely SlERF36 and SlERF39 may involve in BW and drought responses. In this study, transgenic plant with altered expression levels of these genes were generated and analyzed for their stress responses and the involvement regulatory mechanism. The result show that SlERF36 was a functional transcriptional repressor. Overexpression of SlERF36 in transgenic tobacco plants demonstrated that SlERF36 increased susceptibility to BW, decreased osmotic and water tolerance. It is hypothesized that SlERF36 may reduce the expression of defense response and water tolerance response by suppressing the expression of defense response and drought tolerant activators. SlERF35 also function as a transcriptional repressor. Overexpression of SlERF39 cause increased susceptibility to BW and decreased in water stress tolerance, increased expression of ethylene (ET) and jasmonic acid (JA) biosynthesis genes. It is hypothesized that SlERF39 re-depressed the ET and JA signaling by suppressing the expression of certain repressors. Reduced expression of SA led to susceptibility to BW. SlERF39 may reduce the expression of water tolerance response by suppressing the expression of drought tolerant activators but increased the expression of salt tolerant through suppression of salt tolerant suppressor. The transgenic plants generated in this study can be used for future analyses to gain further insight into disease resistant and abiotic cross-talk mechanism of these proteins.

口試委員會審定書 i
誌謝 ii
中文摘要 iii
ABSTRACT iv
縮寫與全名對照表 v
目錄 vii
圖目錄 ix
附錄目錄 x
第一章 前言 1
植物病害防禦機制 1
植物病害與非生物逆境之交互作用 1
青枯病 (Bacterial wilt, BW) 之介紹 2
青枯病反應相關研究 2
AP2 (APETALA2)/ERF 植物乙烯轉錄因子超級家族 3
研究動機 4
第二章 材料與方法 6
1. 植物材料簡介 6
2. 基因選殖常用實驗 6
3. 轉錄活性分析法Transactivation assay 9
4. 番茄以真空吸引浸透法 (Vacuum-infiltration) 接種青枯病菌、細菌性斑點病、細菌性黑腐病及軟腐病之樣品製備 10
5. 檢測基因表現 11
6. 植物轉殖 12
7. 植物之逆境檢測 14
第三章 結果 18
1. 利用生物資訊分析番茄SlERF36與SlERF39基因結構組成及其序列比對分析 18
2. SlERF36與SlERF39 調控轉錄活性分析 (Transactivation assay) 18
3. 35S::SlERF36轉殖菸草之性狀分析 19
4. SlERF36-RNAi番茄轉殖株性狀分析 20
5. 35S::SlERF39轉殖菸草之性狀分析 21
6. SlERF39-RNAi番茄轉殖株性狀分析 23
第四章 討論 25
1. SlERF36與SlERF39為抑制子型 (repressor-type) 轉錄因子 25
2. SlRF36與SlERF39可能負調控植物抗青枯病反應,但可能未參與細菌性軟腐病害反應 25
3. SlRF36與SlERF39負調控植物乾旱抗性,也參與ABA與滲透壓逆境,但可能未參與鹽害反應 27
4. pCAMBIA1301之空載體番茄轉殖株發病異常 28
5. 總結 28
參考文獻 30


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