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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 楊健志 | |
| dc.contributor.author | Ya-Jhen Chen | en |
| dc.contributor.author | 陳亞甄 | zh_TW |
| dc.date.accessioned | 2021-05-19T17:55:30Z | - |
| dc.date.available | 2021-08-22 | |
| dc.date.available | 2021-05-19T17:55:30Z | - |
| dc.date.copyright | 2016-09-08 | |
| dc.date.issued | 2016 | |
| dc.date.submitted | 2016-08-22 | |
| dc.identifier.citation | Aguilera, A. (2002). The connection between transcription and genomic instability. The EMBO Journal 21, 195-201.
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Yu, K., Chedin, F., Hsieh, C.-L., Wilson, T.E., and Lieber, M.R. (2003). R-loops at immunoglobulin class switch regions in the chromosomes of stimulated B cells. Nat Immunol 4, 442-451. Yüce Ö, West SC. (2013). Senataxin, defective in the neurodegenerative disorder ataxia damage response. Molecular and Cellular Biology 33: 406–417.with oculomotor apraxia 2, lies at the interface of transcription and the DNA 謝閔翔. (2015).阿拉伯芥中可能形成 R-loop 基因之探討. 台灣大學碩士論文 | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/7839 | - |
| dc.description.abstract | R-loop為RNA與互補DNA雜合後使另一股DNA分離而單獨存在所形成的三股核酸結構。R-loop的結構包含由轉錄後的RNA與模板DNA (template strand DNA ) 重新配對，及單股的非模板DNA (non-template strand DNA)。植物中第一個R-loop有關之研究，是COOLAIR基因形成R-loop後，失去抑制FLC的作用，而共同調控植物開花。為了深入了解 R-loop是否在植物扮演其他角色，本實驗室謝閔翔與生物產業機電工程系陳倩瑜老師實驗室合作，使用程式語言「C++」來設計出可以搜尋出可能形成R-loop的基因的程式，篩選出了在53個在阿拉伯芥中可能形成R-loop的基因。謝 (2015)已研究LBD18在in vitro及in vivo形成R-loop。本實驗挑選13個基因以胞外轉錄及三種gel mobility shift assay進行研究。13個基因中，部分出現in vitro R-loop形成之特徵性色帶模糊。但同一種基因中，三種gel mobility shift assay得到的實驗結果不盡相同，且LBD18在三種方法中都沒有呈現色帶模糊。另外，本研究發現之前使用urea於染色體DNA抽取可能有缺點，因此使用改良的抽取方法，唯此方法未能再現LBD18在native bisulfite sequencing assay之結果。 | zh_TW |
| dc.description.abstract | R-loop is a triple helix structure consisting of a DNA/RNA hybrid plus a single strand DNA. R-loops are formed when newly synthesized RNA threads back to anneal with the template strand DNA, leaving the non-template strand DNA remaining as a single strand. In Arabidopsis, the expression level of COOLAIR, the first gene discovered that can form R-loops in plants, coordinates flowering with FLC. To explore if R-loop formation is widespread in plants, Min-Hsiang Hsieh designed an R-loop searching program written using C++ together with Dr. Chien-Yu Chen (Department of Bio-industrial Mechatronics Engineering of National Taiwan University) .Fifty-three genes were predicted to be able to form R-loops by the program. Hsieh (2015) found that LOB domain-containing protein 18 (LBD18) form DNA/RNA hybrids in vitro and in vivo. In this study, 13 genes were tested by in vitro transcription and three kinds of gel mobility shift assay. Part of the 13 genes had the distinctive smear of DNA/RNA hybrid in vitro, but different protocol caused different consequences in some genes. In this study, LBD18 did not have smear pattern by three kinds of gel mobility shift assay. Moreover, there was shortcoming of using urea to extract genomic DNA by Hsieh. (2015). Modified genomic DNA extraction protocol did not repeat the native bisulfite sequencing result of LBD18. | en |
| dc.description.provenance | Made available in DSpace on 2021-05-19T17:55:30Z (GMT). No. of bitstreams: 1 ntu-105-R02b22063-1.pdf: 3962271 bytes, checksum: 00273dd870b1588468ff2fe735a168b7 (MD5) Previous issue date: 2016 | en |
| dc.description.tableofcontents | 目錄 1
摘要 3 Abstract 4 縮寫表 5 第一章 緒論 6 1.1 細胞中 DNA/RNA雜合體的生成 6 1.1.1 R-loop生成的特徵 7 1.2 R-loop的存在與基因不穩定性( genomic instability) 8 1.3 調控R-loop的生理機制 9 1.3.1細胞中移除R-loop結構的方法 10 1.3.2防止R-loop形成的生理機制 11 1.4 R-loop的生理功能 13 1.5 R-loop與非編碼RNA (non-codng RNA, ncRNA) 13 1.7依R-loop的序列特徵建立計量模型 14 1.8 建立R-loop基因搜尋程式 15 1.9本篇論文之研究起源與動機 15 第二章 材料與方法 17 2.1植物材料及生長條件 17 2.1.1 植物材料 17 2.1.2植物阿拉伯芥培養基1/2 MS 17 2.1.3種子表面消毒及低溫處理 17 2.1.4種子之無菌培養 18 2.2 The gel mobility shift assay 18 2.2.1 以Phusion DNA polymerase 進行聚合酶連鎖反應 (polymerase chain reaction, RCP) 18 2.2.2 DNA瓊脂糖膠體電泳 (Agarose Gel Electrophoresis) 19 2.2.3 PCR產物之分離與純化 19 2.2.4 TA cloning 20 2.2.5大腸桿菌培養基 20 2.2.6大腸桿菌熱衝擊轉形(Heat-shock Transformation) 21 2.2.7菌落PCR檢定法 21 2.2.8.1利用實驗套組進行質體DNA小量製備 21 2.2.8.2以鹼裂解法進行質體DNA小量製備 22 2.2.9胞外轉錄 (in vitro transcription)方法一 22 2.2.10胞外轉錄方法二 23 2.2.11 胞外轉錄方法三 24 第三章 結果與討論 25 3.1利用R-loop searching program 尋找阿拉伯芥中可能形成R-loop的基因 25 3.1.1 挑選目標基因的條件 26 3.2 以 Gel mobility sift assay 檢驗在in vitro條件下是否有R-loop結構存在 27 3.3 13個候選基因進行GMSA 29 第四章 未來展望 37 參考文獻 39 Figures 44 Table 75 Appendices 76 問答集 95 | |
| dc.language.iso | zh-TW | |
| dc.title | 利用電泳分析程式預測之阿拉伯芥中可能形成
R-loop結構之基因 | zh_TW |
| dc.title | Analysis of the program-predicted R-loop forming genes in Arabidopsis by gel mobility shift assay | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 104-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 黃楓婷 | |
| dc.contributor.oralexamcommittee | 王愛玉,廖憶純,陳佩燁 | |
| dc.subject.keyword | R-loop,阿拉伯芥,電泳分析,基因組DNA抽取, | zh_TW |
| dc.subject.keyword | R-loop,Arabidopsis,gel mobility shift assay,genomic DNA extraction, | en |
| dc.relation.page | 97 | |
| dc.identifier.doi | 10.6342/NTU201603524 | |
| dc.rights.note | 同意授權(全球公開) | |
| dc.date.accepted | 2016-08-22 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科技學系 | zh_TW |
| 顯示於系所單位： | 生化科技學系 | |
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