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研究生:張毓安
研究生(外文):Yu-An Chang
論文名稱:瘤野螟咬食誘導水稻蔗糖轉運蛋白OsSUT4基因表現之調控
論文名稱(外文):Regulation of sucrose transporter gene OsSUT4 in response to rice leaffolder infestation in rice
指導教授:王淑珍王淑珍引用關係
指導教授(外文):Shu-Jen Wang
口試委員:洪傳揚張孟基謝旭亮郭瑋君
口試委員(外文):Chwan-Yang HongMen-Chi ChangHsu-Liang HsiehWoei-Jiun Guo
口試日期:2015-07-31
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:農藝學研究所
學門:農業科學學門
學類:一般農業學類
論文種類:學術論文
論文出版年:2015
畢業學年度:103
語文別:中文
論文頁數:55
中文關鍵詞:水稻蔗糖轉運蛋白瘤野螟
外文關鍵詞:rice (Oryza sativa L.)sucrose transporterrice leaffolder
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植物在生長發育過程中易發生機械性傷害及蟲害,而植物在受到傷害刺激後會引發一連串生理及代謝反應進行防禦或修護。蔗糖是光合作用之主要產物,參與在植物許多代謝路徑中,且醣類在植物受傷害刺激時具有訊息傳遞功能。瘤野螟 [Cnaphalocrocis medinalis (Guenée)] 為水稻主要啃咬型害蟲之一,其習性為捲葉並啃食上表皮及葉肉。本研究利用瘤野螟四齡幼蟲進行試驗,發現受蟲咬的葉片蔗糖含量會降低;正在發育的未完全展開葉蔗糖含量則會增加,推測水稻植株在瘤野螟咬食後會傾向將蔗糖從受傷部位轉移到正在發育的部位。蔗糖轉運蛋白 (sucrose transporter, SUT) 為植株中負責將蔗糖裝載及卸載於韌皮部的主要蛋白,以助蔗糖於不同器官間進行長距離運輸。水稻SUT基因家族中之OsSUT4基因表現量會受到瘤野螟侵咬刺激而明顯提升,瘤野螟咬食後植株不同葉位OsSUT4不會有系統型的反應,但根部OsSUT4表現將下降。進一步利用訊息傳導因子相關基因及生合成抑制劑探討此誘導表現的機制,發現茉莉酸 (jasmonic acid, JA)、水楊酸 (salicylic acid, SA)、離層酸 (abscisic acid, ABA)、乙烯 (ethylene) 會受到蟲咬刺激而產生,且在傷口處會有過氧化氫 (hydrogen peroxide, H2O2) 的累積。而以茉莉酸、離層酸及過氧化氫生合成抑制劑處理後會降低蟲咬誘導OsSUT4基因表現之反應,顯示這些因子會參與在蟲咬誘導OsSUT4基因表現的路徑中。

Plants were often suffered from many biotic and abiotic stresses such as mechanical wounding and insect herbivores. Plants protect themselves by several defense responses. Sucrose is the major product of photosynthesis. It is involved in many metabolic regulatory pathways and plays an important role in the plant defense responses as signaling molecule. Rice leaffolder [Cnaphalocrocis medinalis (Guenée)] is one of chewing insects. It attacks rice plants by folding leaves and scraping the mesophyll tissues. In this study, we found that after fourth-instar larvae feeding, sucrose contents were decreased in the leaf damaged by leaffolder but increased in developing leaf. Thus, it was suggested that sucrose translocation from damaged leaf to developing sink tissues may be promoted in leaffolder-attacked rice plants. Sucrose transporters (SUT) is the key factor in charge of sucrose loading and unloading in phloem for long-distance translocation in plants. Gene expressions of OsSUT4, a rice SUT gene family member, were significantly induced in leaffolder infestated leaves, but the changes of transcript levels in other leaves were not significant. Rice leaffolder infestation induced jasmonic acid, salicylic acid, abscisic acid and ethylene biosynthesis gene expressions in rice seedlings. Hydrogen peroxide were also accumulated in wounded leaves. Furthermore, leaffolder-induced OsSUT4 gene expressions in leaf tissues would be repressed by inhibitors of jasmonic acid, abscisic acid and hydrogen peroxide biosynthesis. Taken together, the aforementioned results suggested that OsSUT4 gene expression induced by leaffolder infestation was mediated jasmonic acid, abscisic acid or hydrogen peroxide as signalings.

目錄
誌謝…………………………………………………………………………………i
中文摘要…………………………………………………………………………ii
英文摘要.………………………………………………………………………iii
目錄……………………………………………………………………………………v
表目錄………………………………………………………………………………ix
圖目錄………………………………………………………………………………ix
附表及附圖目錄……………………………………………………………x
縮寫字對照………………………………………………………………………xi
前言…………….……………………………………………………………………1
1. 植物面對啃咬型昆蟲之防禦策略……………………1
2. 瘤野螟對水稻的影響……………………………………………2
3. 醣類分子於植物面對機械性傷害時扮演之角色…………………3
4. 水稻蔗糖轉運蛋白之分類及功能…………………………………………3
5. 蔗糖轉運蛋白於機械性傷害與昆蟲侵害下之反應…………………………5
6. 植物受機械性傷害之訊息調控路徑………………………………………6
7. 本論文之研究主題……………………………………………………………9
材料與方法…….……………………………………………………………………11
1. 實驗材料.……………………………………………………………………11
2. 種子消毒與植物種植.…………………………………………………11
3. 材料處理.……………………………………………………………………11
3.1瘤野螟餵養處理………………………………………………………11
3.2水稻植株之荷爾蒙處理………………………………………………11
3.3水稻植株之荷爾蒙抑制劑及過氧化氫生合成抑制劑處理………12
4. 基因表現分析…………………………………………………………12
4.1 總RNA萃取………………………………………………………12
4.2 TURBO DNase 處理………………………………………………13
4.3 RNA電泳…………………………………………………………13
4.4 即時反轉錄聚合酶連鎖反應 (real-time RT-PCR)………………13
5. 過氧化氫(H2O2)定性及定量測定………………………………………14
5.1 瘤野螟咬食後之過氧化氫定性觀察…………………………………14
5.2瘤野螟咬食後植株體內過氧化氫含量變化…………………………14
6. 總可溶性醣類含量測定……………………………………………………15
7. 葡萄糖與蔗糖含量測定……………………………………………………15
7.1可溶性醣類萃取…………………………………………………………15
7.2葡萄糖含量測定…………………………………………………………16
7.3蔗糖含量測定……………………………………………………………16
結果…………………………………………………………………………………18
1. 瘤野螟咬食後,植株不同部位醣類含量的變化…………………………18
2. 瘤野螟咬食對水稻植株OsSUTs基因表現之影響………………………18
3. 瘤野螟咬食後,水稻植株不同部位的OsSUTs基因表現情形…………19
4. 探討荷爾蒙因子在瘤野螟誘導OsSUT4基因表現之調控………………19
4.1 茉莉酸對瘤野螟誘導OsSUT4基因表現的影響……………………19
4.2 離層酸對瘤野螟誘導OsSUT4基因表現的影響…………………20
4.3 水楊酸對瘤野螟誘導OsSUT4基因表現的關係…………………20
4.4 乙烯對瘤野螟誘導OsSUT4基因表現的關係………………………21
5. 過氧化氫在瘤野螟誘導OsSUT4基因表現之調控………………………22
討論……………………………………………………………………………………23
1. 蔗糖轉運蛋白在瘤野螟咬食後的差異性表現……………………………23
2. 瘤野螟咬食後各訊息因子影響OsSUT4基因表現之探討……………23
3. 機械性傷害與啃咬型昆蟲對OsSUT4基因調控之異同…………………25
4. 比較不同食性昆蟲對OsSUTs影響之異同………………………………26
5. OsSUT4在水稻植株受到瘤野螟咬食後可能扮演之角色……………27
6. 結語與未來展望…………………………………………………………28
參考文獻……………………………………………………………………………38
圖目錄
圖一、瘤野螟處理下,水稻植株中不同部位醣類物質含量的變化……30
圖二、瘤野螟處理對水稻植株OsSUTs基因表現之影響……………………31
圖三、瘤野螟處理對水稻植株不同部位OsSUTs基因表現之影響……………32
圖四、茉莉酸對瘤野螟誘導OsSUT4基因表現之影響……………………33
圖五、離層酸對瘤野螟誘導OsSUT4基因表現之影響……………………34
圖六、水楊酸對瘤野螟誘導OsSUT4基因表現之影響……………………35
圖七、乙烯對瘤野螟誘導OsSUT4基因表現之影響…………………………36
圖八、過氧化氫對瘤野螟誘導OsSUT4基因表現之影響………………………37
附表目錄
附表一、木村式水耕液配方………………………………………………53
附表二、Real-time RT-PCR之專一性引子………………54

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