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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林讚標 | |
| dc.contributor.author | Chien-Yu Lu | en |
| dc.contributor.author | 魯建宇 | zh_TW |
| dc.date.accessioned | 2021-06-13T01:37:31Z | - |
| dc.date.available | 2016-08-09 | |
| dc.date.copyright | 2011-08-09 | |
| dc.date.issued | 2011 | |
| dc.date.submitted | 2011-08-02 | |
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Proc. Natl. Acad. Sci. 101: 9508-9512. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/30118 | - |
| dc.description.abstract | 植物為了選擇一個最合適的發芽時期，透過複雜的機制來調控種子休眠，近幾年來關於休眠方面的研究雖然很多，但是關於休眠的分子調控機制仍然了解得不夠透徹，尤其是關於非模式植物方面的研究了解的更少。因此，本研究以山櫻花種子做為材料，研究休眠種子在打破休眠時的相關基因調控。山櫻花新鮮收穫的種子具有很強的休眠性，需要暖溫處理和隨後的低溫層積處理才能有效的打破休眠，研究顯示，山櫻花種子需要四週的暖溫層積，和隨後八週的低溫層積，才會獲得最高的發芽率，此特殊的機制是阿拉伯芥所不具有的。低溫層積對休眠種子所造成的影響，具有何種生理功能以及基因的調控已有許多的研究；但暖溫層積有何種功效目前還不太清楚，因此將研究重點放在暖溫層積方面，利用分子生物學的抑制性扣減雜交技術，尋找在暖溫層積過程中休眠種子的基因調控模式，探討暖溫層積對於休眠種子的影響。將成熟種子採收洗淨並室內陰乾三天，之後直接進行層積處理，暖溫層積處理在進入第二和第七天分別進行扣減抑制雜合反應，完成兩天和七天暖溫層積處理之扣減式基因庫，先用Reverse Northern確認所篩選到的基因是否確實有差異性表現，再以電泳方式除去基因大小相同的片段後，才將其送去定序。序列分析比對後，成功得到暖溫層積後被大量誘導表現以及被抑制的基因共59個，分別在暖溫層積過程中扮演著各自的生理功能，包含代謝、轉錄和轉譯、訊息傳導、細胞中的過程、生長發育、養分儲存、逆境相關蛋白等。暖溫層積過程中ABA反應負調控者(negative regulator) RACK1和PP2CA的大量表現，以及ABA反應正向調控因子(positive component) 1-phosphatidylinositol-4- phosphate 5-kinase的抑制，具有重要的生理意義，顯示暖溫層積或許透過調控這些基因來降低胚軸對ABA的敏感度，以便為將來種子由休眠狀態轉變為種子發芽狀態做好準備。此外，在暖溫層積進入第七天時，山櫻花種子胚軸內ABA含量也有顯著性的下降情況。綜合研究結果來看，雖然在只有暖溫層積的情況下山櫻花種子不能發芽(GAs含量低)，但暖溫層積會透過降低種子胚軸內的ABA含量，以及透過大量表現ABA反應負調控者和抑制ABA反應正向調控因子來減少種子胚軸對ABA的敏感度，來調控種子的休眠解除。 | zh_TW |
| dc.description.abstract | Seed dormancy is regulated by complex networks in order to optimize the timing of germination. Although over 5000 publications on seed dormancy have appeared in the past decades, the underlying molecular regulatory mechanisms are not fully understood. Fresh seeds of Formoan cherry (Prunus campanulata Maxim), a popular ornamental tree, are deeply dormant and require warm stratification followed by cold stratification for complete dormancy-breaking. This operation is special to Formoan cherry but not to Arabidopsis. Many studies have revealed the effect of cold stratification on dormancy-breaking, but the effect of warm stratification is unclear. In this study we used the subtractive suppression hybridization (SSH) to study the gene regulation of prunus seeds undergoing warm stratification. Formoan cherry seeds were harvested, cleaned and air-dried for 3 days to reduce the water content to 15% of fresh weight and followed by warm stratification. At the second and seventh days of warm treatment, seed RNAs were extracted for SSH experiments. After constructed the subtraction libraries of second and seventh days against zero day of warm stratification, we performed reverse northern assays to confirm the upregulated and down-regulated genes. Clones obtained from this study were sequenced after removing duplicated clone containing the same length of DNA fragments. Using the website to blast the fragment sequences, we obtained 59 genes were up-regulated and down-regulated that play functions in metabolism, transcription, translation, signal transduction, growth and development, nutrient storage and stress-related. Interestingly, two ABA negative regulators, RACK1 and PP2CA were highly expressed and one positive component, 1-phosphatidylinositol-4- phosphate 5-kinase was repressed. These three regulators could reduce the ABA sensitivity of prunus seeds in the beginning of warm stratification. Moreover, we found that the ABA content was significantly decreased at the seventh days of warm stratification. Taken together, warm stratification treatment alone can not promote the seed germination due to low GA content, but can reduce the ABA sensitivity by expressing ABA negative regulators and repressing positive components in the beginning when ABA content is still high and followed by reduced the ABA content in the seeds. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T01:37:31Z (GMT). No. of bitstreams: 1 ntu-100-R97b42014-1.pdf: 3588719 bytes, checksum: 1dac44ea1bf0fff94f30e51d40865550 (MD5) Previous issue date: 2011 | en |
| dc.description.tableofcontents | 目錄
致謝----------------------------------------------------------------------------------------------II 摘要----------------------------------------------------------------------------------------------III Abstract------------------------------------------------------------------------------------------IV 第一章 序論---------------------------------------------------------------------------------1 1.1 起源-----------------------------------------------------------------------------------1 1.2 種子休眠的分類--------------------------------------------------------------------1 1.3 植物荷爾蒙與種子休眠-----------------------------------------------------------1 1.3.1激勃素(GAs)與種子休眠解除和發芽相關研究-------------------------3 1.3.2離層酸(ABA)與種子休眠的相關研究------------------------------------3 1.4 解除休眠的相關研究--------------------------------------------------------------4 1.5 抑制性扣減雜交--------------------------------------------------------------------6 1.6 山櫻花種子之休眠及解除機制的研究-----------------------------------------7 1.7 研究策略與目標--------------------------------------------------------------------8 第二章 材料與方法------------------------------------------------------------------------9 2.1 植物材料採集與處理--------------------------------------------------------------9 2.2 暖溫層積處理-----------------------------------------------------------------------9 2.3 山櫻花種子的水分含量與百粒重-----------------------------------------------9 2.4 扣減抑制雜合反應-----------------------------------------------------------------9 2.5 Reverse Northern blot analysis-----------------------------------------------------9 2.6 資料庫鑑定比對-------------------------------------------------------------------10 2.7 Real-time PCR-----------------------------------------------------------------------10 2.8 利用ELISA定量種子胚軸內ABA含量--------------------------------------10 第三章 結果--------------------------------------------------------------------------------11 3.1 山櫻花種子的構造和縱剖面-----------------------------------------------------11 3.2 暖溫層積處理對山櫻花種子水分含量與百粒重的影響--------------------11 3.3 暖溫層積處理過程中山櫻花種子胚軸ABA含量的變化------------------12 3.4 暖溫層積處理過程中山櫻花休眠種子的基因變化--------------------------12 3.4.1 兩天暖溫層積處理後表現量下降的基因-------------------------------12 3.4.2 七天暖溫層積處理後表現量下降的基因-------------------------------14 3.4.3 兩天暖溫層積處理後表現量上升的基因-------------------------------18 3.4.4 七天暖溫層積處理後表現量上升的基因-------------------------------20 3.5以Reverse Northern blot和Real time技術再次確認基因變化------------25 第四章 討論---------------------------------------------------------------------------------26 4.1暖溫層積處理對於山櫻花休眠種子之影響------------------------------------26 4.2暖溫層積處理過程中山櫻花休眠種子的基因調控---------------------------26 4.2.1暖溫層積處理後表現量下降的基因--------------------------------------27 4.2.2暖溫層積處理後表現量上升的基因--------------------------------------27 4.3暖溫層積處理對於山櫻花種子之休眠及解除機制探討--------------------28 4.4未來研究方向-----------------------------------------------------------------------30 參考文獻----------------------------------------------------------------------------------------31 圖表----------------------------------------------------------------------------------------------40 表格----------------------------------------------------------------------------------------------48 附錄----------------------------------------------------------------------------------------------53 圖目錄 Fig. 1 Fruits and seeds of Formoan cherry-------------------------------------------------40 Fig. 2 Moisture content and 100 seeds weight of Formoan cherry seeds--------------41 Fig. 3 Abscisic acid content of embryonic axis from fresh or warm stratified Formoan cherry seeds------------------------------------------------------------------42 Fig. 4 Selection of differentially expressed genes during warm stratification of Formoan cherry seeds------------------------------------------------------------------43 Fig. 5 Screening for differential expression genes after 2-day warm stratification with reversed northern blot analysis-------------------------------------------------44 Fig. 6 Screening for differential expression genes after 7-day warm stratification with reversed northern blot analysis-------------------------------------------------45 Fig. 7 Screening for differential expression genes after 7-day warm stratification with reversed northern blot analysis-------------------------------------------------46 Fig. 8 The relative expression level of RACK1B and PP2CA under warm stratification-----------------------------------------------------------------------------47 表目錄 表1. 山櫻花新鮮種子兩天暖溫層積處理後表現量下降的基因---------------------48 表2. 山櫻花新鮮種子七天暖溫層積處理後表現量下降的基因---------------------49 表3. 山櫻花新鮮種子兩天暖溫層積處理後表現量上升的基因---------------------50 表4. 山櫻花新鮮種子七天暖溫層積處理後表現量上升的基因---------------------51 | |
| dc.language.iso | zh-TW | |
| dc.subject | 離層酸 | zh_TW |
| dc.subject | 扣減抑制雜合反應 | zh_TW |
| dc.subject | 暖溫層積 | zh_TW |
| dc.subject | 休眠種子 | zh_TW |
| dc.subject | 山櫻花 | zh_TW |
| dc.subject | warm stratification | en |
| dc.subject | abscisic acid | en |
| dc.subject | Formoan cherry | en |
| dc.subject | seed dormancy | en |
| dc.subject | subtractive suppression hybridization | en |
| dc.title | 暖溫層積處理過程中山櫻花休眠種子的基因調控 | zh_TW |
| dc.title | The gene regulated in dormant seed of Prunus campanulata during warm stratification | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 99-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 張英?,鄭萬興,靳宗洛,簡慶德 | |
| dc.subject.keyword | 山櫻花,休眠種子,暖溫層積,離層酸,扣減抑制雜合反應, | zh_TW |
| dc.subject.keyword | Formoan cherry,seed dormancy,warm stratification,abscisic acid,subtractive suppression hybridization, | en |
| dc.relation.page | 76 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2011-08-02 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 植物科學研究所 | zh_TW |
| 顯示於系所單位： | 植物科學研究所 | |
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