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研究生:曾冠儒
研究生(外文):Guan-RuTseng
論文名稱:探討不同品系阿拉伯芥根部受砷調控轉錄體之分析
論文名稱(外文):Ecotypic variation in genome-wide transcription profiles induced by arsenic in Arabidopsis roots
指導教授:黃浩仁
指導教授(外文):Hao-Jen Huang
學位類別:碩士
校院名稱:國立成功大學
系所名稱:生命科學系碩博士班
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:2010
畢業學年度:98
語文別:中文
論文頁數:73
中文關鍵詞:阿拉伯芥微陣列
外文關鍵詞:Arabidopsisarsenicmicroarray
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砷(Arsenic)為有毒性的類金屬,常以正五價或正三價無機鹽的型態廣泛存在自然界中。在地下水污染的環境,砷會透過植物進入食物鏈,進而對人體造成致癌危險性。然而目前對砷影響植物生長之分子機制尚未明瞭,因此本篇研究探討當植物受到砷(V)逆境時,如何在分子層次調控轉錄體之變化以面對此種非生物性逆境。首先,我們以砷(V)處裡不同品系阿拉伯芥野生型Col-0與野生型Ws-2 兩天,則會發現Col-0根部生長顯著較Ws-2長。另外,深入測定累積砷的含量,發現以100 μM砷(V)處裡三小時後,Ws-2累積砷元素的量顯著高過Col-0所累積的1.86倍。綜合上述實驗,Col-0相對於Ws-2展現出耐砷的表現型。接著利用ATH1 基因晶片,進行核酸微陣列(DNA microarray)分析Col-0與Ws-2根部受砷調控轉錄體。在以100 μM砷(V)處理短時間(1.5+3 h)後,在Col-0與Ws-2中均顯示轉運蛋白家族 Aquaporin與LeOPT1-like基因表現量下降數目相對最多且一致性降低。Ws-2則在負責解毒作用的GST和GST-Tau與ABC家族轉運蛋白會誘發上升較多基因。在定義為耐受性相關基因類群中可分析出14個轉錄因子分屬於AP2/EREBP、bHLH、C2H2、C3H、MBF1、MYB-related、Trihelix、WRKY與ZIM; 而Ethylene荷爾蒙相關的基因僅在Col-0會受調控,顯示Ethylene可能參與Col-0面對砷(V)逆境的訊息傳遞。綜合以上,本篇研究結果為非常有價值的資源庫,提供方向探討植物對砷(V)逆境的耐受性機制與這些機制如何在基因轉錄的層次受到調控。
Arsenic (As) is considered as the most common toxic metalloid which is widely found in the environment. It primarily exists in the form of inorganic arsenate or arsenite. Under the circumstances of contaminated groundwater, As would penetrate into the food chain through irrigation of vegetables and crop plants and then threatens human health to cancer. However, what impact that As causes to the molecular response and gene expression of plants has not been extensively characterized currently, therefore this study aims to explore how plants respond the nature of toxicity and the mechanism of signal transductions when facing the abiotic stress. First, when Col-0 and Ws-2 seedlings were subjected to arsenate treatment for 2 days, the root elongation rate of Col-0 was found significantly higher than that of Ws-2. In addition, after the exogenous treatment of 100 μM arsenate for 3 hours, the result showed that arsenic accumulation in Ws-2 was 1.86 times higher in comparing with Col-0’s. Accordingly, Col-0 exhibited more tolerance to arsenate stress than Ws-2. Next, the ATH1 gene chip was used to compare the transcriptome of Col-0 and Ws-2. With the treatment of 100 μM arsenate for a short period of time (1.5hrs and 3 hrs), Aquaporin transporter family and LeOPT1-like transporter family genes showed more down-regulated gene numbers and were repressed with consistency. Besides, genes encoding glutathione transferase (GST) and ABC transporter were found to be significantly induced in Ws-2. Based on the definition of the tolerance-associated gene, 14 transcription factor genes could be sorted to 9 families : AP2/EREBP, bHLH, C2H2, C3H, MBF1, MYB-related, Trihelix, WRKY and ZIM. And Ethylene-related genes were found only regulated in Col-0. This might suggest that when Col-0 face the arsenate stress, Ethylene involved in the process. To sum up, this study presents a comprehensive survey of global transcriptional regulation under arsenate stress. The results described here will help to further our understanding of the underlying mechanisms of arsenate toxicity and tolerance in plants.
摘要 2
Abstract 3
1.前言 10
1.1重金屬與類金屬的毒害 10
1.2砷的介紹 10
1.3砷的吸收 11
1.4砷的毒害 11
1.5砷的解毒作用 12
1.6研究目的 13
2.材料與方法 14
2.1阿拉伯芥植株 14
2.2阿拉伯芥植株的培養 14
2.3根長實驗 14
2.4利用感應耦合電漿原子發射光譜分析儀測定根部細胞砷含量 15
2.5 RNA的萃取 16
2.6反轉錄聚合?反應 (RT-PCR) 17
2.7聚合?鏈鎖反應 (PCR) 17
2.8微陣列實驗 18
2.9微陣列數據分析 18
3.結果 19
3.1 在五價砷(V)逆境下, 阿拉伯芥野生型Col-0較野生型Ws-2展現出較高的耐受性 19
3.2 阿拉伯芥野生型Ws-2會在砷(V)逆境中累積大量的砷元素 20
3.3 利用微陣列(Microarray)探討不同品系阿拉伯芥在砷(V)處理之基因轉錄 20
3.4 在正常生長條件下,會在Col-0或Ws-2相對表現量較高的基因 21
3.5 阿拉伯芥在砷(V)逆境下所受調控的基因群 22
3.6 耐受性相關的基因 (Tolerance-associated genes) 24
3.7 阿拉伯芥在砷(V)逆境中受調控的荷爾蒙相關基因 26
4.討論 27
4.1阿拉伯芥在短時間砷(V)逆境下所受調控的基因群 27
4.2比較不同品系阿拉伯芥在短時間砷(V)逆境下所受調控的基因群 28
5.參考文獻 31
6.自述 73
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