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姓名 曾梓維(Tzu-wei Tseng)  查詢紙本館藏   畢業系所 生命科學系
論文名稱 探討阿拉伯芥兩個MYB-related轉錄因子在糖訊息傳遞中所扮演的角色
(Two MYB-related transcription factors play opposite roles in sugar signaling in Arabidopsis)
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摘要(中) MYB蛋白質是一群具有MYB保守性區段的轉錄因子,此DNA結合單元由一至三個各約51個胺基酸的不完全重複組成。植物MYB家族可依DNA結合單元中所含的不完全重複數目分為三個次家族,含有R1、R2、R3三個MYB重複片段屬於R1R2R3 次家族,含有R2、R3兩個MYB重複片段屬於R2R3,而含有一個MYB重複片段則為MYB-related次家族。先前研究中已證明水稻中三個MYB-related轉錄因子,OsMYBS1、OsMYBS2和OsMYBS3,會參與在糖訊息與賀爾蒙訊息調控α-澱粉降解酶基因表現的途徑中。為了更進一步了解這一群MYBS蛋白質在阿拉伯芥中的生理功能,我們利用水稻OsMYBSs蛋白質之胺基酸序列,找出阿拉伯芥中四個胺基酸序列相似的MYBS蛋白質,命名為AtMYBS1、AtMYBS2、AtMYBS3及AtMYBS5。本篇論文進行AtMYBS1及AtMYBS2蛋白質在細胞中的位置分析,結果顯示AtMYBS1和AtMYBS2位於細胞核內。在外表型分析中,我們發現atmybs1/s2雙重突變株的葉、莖以及花苞都呈現淺綠色的外觀相較於野生型植株、atmybs1突變株與atmybs2突變株,再進一步分析葉綠素含量及葉綠素生合成基因的表現量,也顯示atmybs1/s2雙重突變株的葉綠素含量及葉綠體MEP pathway基因的表現量都較野生型植株低。在種子萌芽以及早期幼苗發育階段,atmybs1突變株對於4% 和6% 葡萄糖以及0.5 μM, 1 μM, 3 μM 和5 μM 離層酸具有高敏感性,atmybs2突變株則是對於4% 和6% 葡萄糖以及0.5 μM, 1 μM, 3 μM 和5 μM 離層酸有低敏感性,而atmybs1/s2雙重突變株的敏感性介於atmybs1和atmybs2突變株之間。這些結果指出種子萌芽階段,AtMYBS1可能扮演著促進種子萌芽及幼苗生長的角色,而AtMYBS2扮演著抑制種子萌芽及幼苗生長的角色,並參與調控糖訊息傳遞途徑以及影響葉綠素的生合成。
摘要(英) MYB proteins are transcription factors containing around 1-3 MYB DNA-binding domains which are composed of 51 amino acids. Base on repeat number of the MYB DNA-binding domain, MYB proteins in plant have been classified into three subfamilies, R1R2R3, R2R3 and R1/2. Previously, three R1/2 MYB transcription factors, OsMYBS1, OsMYBS2 and OsMYBS3 have been demonstrated that are involved in sugar- and hormone-regulated α-amylase gene expression in rice. To study the physiological functions of MYBS proteins in Arabidopsis, we identified four OsMYBS homologous genes, AtMYBS1, AtMYBS2, AtMYBS3 and AtMYBS5. The subcellular localization analysis showed that the AtMYBS1 and AtMYBS2 were localized in nucleus. The mutant of AtMYBS1, atmybs1, exhibited glucose and abscisic acid hypersensitivity phenotype during seed germination and early seedling growth stage;whereas the mutant of AtMYBS2, atmybs2 exhibited hyposensitive to glucose and ABA. The atmybs1/s2 double mutant showed intermediate phenotype for glucose and ABA response between those of the single mutants. These results indicated that the AtMYBS1 plays as a positive regulator and the AtMYBS2 is a negative regulator during seed germination and early seedling growth stage. Compared to wild type, atmybs1 mutant and atmybs2 mutant, the atmybs1/s2 double mutant plants showed pale green leaves, stems and floral buds phenotype. The level of chlorophyll and transcripts of chlorophyll biosynthesis related genes were lower in atmybs1/s2 double mutant than wild type. We therefore conclude that AtMYBS1 and AtMYBS2 are involved in sugar signaling and chlorophyll biosynthesis in Arabidopsis.
關鍵字(中) ★ 葉綠素生合成
★ 離層酸
★ 糖
關鍵字(英) ★ ABA
★ chlorophyll biosynthesis
★ MYBS
★ sugar
論文目次 中文摘要..........................................I
Abstract.........................................II
縮寫檢索表......................................III
本文目錄..........................................V
表目錄..........................................VII
圖目錄..........................................VII
附表目錄.......................................VIII
附圖目錄.......................................VIII
本文目錄
一、緒論...............................................1
1.植物糖訊息傳遞及基因調控研究.........................1
1-1葡萄糖影響植株生長發育..............................1
1-2植物對糖的感應及訊息傳遞............................1
1-3糖與植物荷爾蒙間訊息傳遞途徑........................2
1-4糖訊息與離層酸訊息之對話............................3
2.MYB轉錄因子之介紹....................................4
2-1 分類與功能介紹.....................................5
2-2 水稻OsMYBSs參與糖訊息傳遞途徑......................6
2-3 阿拉伯芥AtMYBSs參與在糖訊息調控的短暫性分析........7
二、研究目的...........................................8
三、材料與方法.........................................9
Plant Materials........................................9
Primers................................................9
Isolation of AtMYBSs cDNA..............................9
Plasmids..............................................10
RT-PCR................................................10
Real-time RT-PCR......................................11
Analysis of AtMYBSs Localization......................11
Analysis of Chlorophyll and Carotenoid Contents.......11
Germination, Cotyledon Greening and True Leaf Expansion Analysis..............................................12
Agrobacterium-mediated gene transformation in Arabidopsis
......................................................12
四、實驗結果..........................................13
1.阿拉伯芥AtMYBSs 之特性分析..........................13
1-1 R1/2 MYB轉錄因子在水稻與阿拉伯芥間的演化樹關係圖..13
1-2 AtMYBS1和AtMYBS2之定位分析........................14
2.阿拉伯芥atmybs突變株之分析..........................14
2-1 atmybs1/s2雙重突變株之篩選........................15
2-2偵測AtMYBS1與AtMYBS2基因表現的程度.................15
2-3阿拉伯芥atmybs1/s2雙重突變株之外觀分析.............16
2-4葉綠素及類胡蘿蔔素含量之分析.......................17
2-5葉綠素生合成途徑MEP pathway之分析..................17
3.阿拉伯芥AtMYBSs參與在糖訊息調控的外觀分析...........18
3-1 黑暗處理下AtMYBSs表現的程度.......................18
3-2 高濃度葡萄糖處理下atmybs1、atmybs2突變株以及atmybs1/s2雙重突變株之耐受性分析................................19
3-3 木糖醇培養基為滲透壓控制組........................20
4.阿拉伯芥AtMYBSs參與在離層酸訊息調控的外觀分析.......21
4-1 不同濃度ABA處理下atmybs1、atmybs2突變株以及atmybs1/s2雙重突變株之耐受性分析..................................21
五、討論..............................................23
六、參考文獻..........................................29
參考文獻 吳珮綺,阿拉伯芥AtMYBSs基因參與在糖訊息及離層酸訊息傳遞之研究,中央大學生命科學所碩士論文,民國九十八年。
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指導教授 陸重安(Chung-an Lu) 審核日期 2011-12-1
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