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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 黃鵬林 博士(Dr. Pung-Ling Huang) | |
| dc.contributor.author | Yung-Wei Cheng | en |
| dc.contributor.author | 程永煒 | zh_TW |
| dc.date.accessioned | 2021-06-16T17:35:21Z | - |
| dc.date.available | 2017-08-27 | |
| dc.date.copyright | 2012-08-27 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-08-15 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/64218 | - |
| dc.description.abstract | 篩選標誌基因之表達有助於分離出轉殖細胞，一旦確立轉殖事件後，篩選標誌基因便不具有其他的用途，若持續添加抗生素或殺草劑等篩選試劑，則易降低轉殖細胞增殖及分化的能力。另外，轉殖植株之抗藥性基因可能影響生態環境的平衡，易引發大眾對其安全性的疑慮。本研究利用可受水楊酸誘導之菸草PR-1a 啟動子與轉位酶 (transposase) 基因融合，應用於四種轉位子 (transposon) 系統，以 3 mM 水楊酸處理轉殖蝴蝶蘭癒傷組織及菸草葉圓片5天，接著抽取基因組 DNA 進行聚合酶連鎖反應 (polymerase chain reaction, PCR)，將 PCR 所得到的產物定序分析，結果顯示轉位事件 (transposition) 發生後，產生四種轉位類型，除了預期的左邊界 (left border) 及右邊界 (right border)，也保留了35 - 128 bp之部分質體序列。本研究進一步針對篩選標誌基因轉位後是否再插入其它位置進行探討，以南方氏雜交法進行分析，轉殖 pGcET 和 pGEnT 植株只雜合到轉位後之片段，並未獲得篩選標誌基因片段，顯示轉位作用已達到篩選標誌基因移除之目標。 | zh_TW |
| dc.description.abstract | Expression of selectable marker genes allows scientists to identify and isolate the transgenic cells. Once the transgenic plant has been generated, marker gene generally no longer serves an essential purpose. Furthermore, the propagation and development of transgenic cells will be retarded if the antibiotic or herbicide remains in the growth medium. Besides, it is concerned by publics about biosafety of antibiotic and herbicide resistance genes in transgenic plants. We have developed four different constructs under the control of the promoter PR-1afor inducible transposon systems as marker-free strategy and theses constructs were transformed into Phalaenopsis calli and tobacco plants. Transgenic plants were treated with 3 mM salicylic acid for 5 days to eliminate marker genes. Four patterns of transposition have been identified base on sequence analysis. Residual DNA sequences of 35 - 128 bp were fused as footprints after excision of the transposon. Southern analysis confirmed further that the herbicide resistance geneEPSPS,had been removed together with the transposition. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T17:35:21Z (GMT). No. of bitstreams: 1 ntu-101-R99628132-1.pdf: 2704327 bytes, checksum: 648201204009b1e4c390f20b521bec00 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | 摘要 i
Abstract ii 壹、前言 1 貳、前人研究 2 一、篩選標誌基因 2 (一) 抗生素抗性基因(antibiotics resistance gene) 2 (二) 殺草劑抗性基因(herbicide resistance gene) 3 (三) 正向篩選基因 (Positive selectable gene) 5 二、篩選標誌基因移除策略 (marker free strategies) 6 (一) 共轉型法 (Co-transformation) 7 (二) 定點重組法 (Site-specific recombination) 8 (三) 轉位子系統 (Transposon system) 8 三、應用誘導型啟動子移除篩選標誌基因 9 四、轉位機制及修復模式 11 參、材料與方法 13 一、試驗材料 13 (一) 質體材料 13 (二) 試驗菌種 13 (三) 植物材料 15 二、試驗方法 15 (一) 農桿菌媒介轉殖法 15 (二) 擬轉殖株分析 16 (三) 水楊酸誘導轉位試驗 19 肆、結果 21 一、水楊酸誘導處理之最佳濃度 21 二、聚合酶連鎖反應分析 21 三、轉位後序列分析 32 四、南方氏雜交分析 37 五、轉殖蝴蝶蘭癒傷組織之再生及 GUS活性組織化學染色分析 37 伍、討論 45 一、水楊酸誘導型PR-1a啟動子 45 二、水楊酸誘導轉位子系統發生轉位之現象 46 三、轉位機制及修復模式 46 四、轉位後之再插入 49 陸、結語 51 柒、參考文獻 52 捌、附錄 63 | |
| dc.language.iso | zh-TW | |
| dc.subject | 轉位酶 | zh_TW |
| dc.subject | 篩選標誌基因 | zh_TW |
| dc.subject | 轉位子 | zh_TW |
| dc.subject | Ac末端反向重複序列 | zh_TW |
| dc.subject | selectable marker gene | en |
| dc.subject | transposase | en |
| dc.subject | transposon | en |
| dc.subject | Acterminal inverted repeat | en |
| dc.title | 應用轉位子系統去除轉殖蝴蝶蘭篩選標誌基因之研究 | zh_TW |
| dc.title | Application of Transposon System for Selectable Marker Elimination in Transgenic Phalaenopsis | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 廖芳心 博士(Dr. Fang-Shin Liao),常玉強 博士(Dr. Yuh-Chyang Charng),杜宜殷 博士(Dr. Yi-Yin Do) | |
| dc.subject.keyword | 篩選標誌基因,轉位子,Ac末端反向重複序列,轉位酶, | zh_TW |
| dc.subject.keyword | selectable marker gene,transposon,Acterminal inverted repeat,transposase, | en |
| dc.relation.page | 66 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-08-15 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 園藝學研究所 | zh_TW |
| 顯示於系所單位： | 園藝暨景觀學系 | |
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