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研究生:唐儀倩
研究生(外文):I-chen Tang
論文名稱:以農桿菌轉殖外來的DNA片段作為文心蘭‘GowerRamsey’之品種分子標誌
論文名稱(外文):Agrobacterium-mediated transformation of foreign DNA sequence as moleculer marker in Oncidium ‘Gower Ramsey’
指導教授:高建元高建元引用關係
指導教授(外文):Chien-Yuan kao
學位類別:碩士
校院名稱:國立宜蘭大學
系所名稱:園藝學系碩士班
學門:農業科學學門
學類:園藝學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
中文關鍵詞:文心蘭農桿菌品種
外文關鍵詞:Oncidium ‘Gower Ramsey’PLBAgrobacterium
相關次數:
  • 被引用被引用:2
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  • 收藏至我的研究室書目清單書目收藏:1
本實驗的目的為建立蘭花的基因轉殖系統及建立分子標誌系統來確認所轉殖的片段可以穩定的遺傳到其增殖的子代。因此該品種的子代(實生或分生)可以具體的以PCR,讀出其DNA序列所代表的品種。
本實驗利用農桿菌進行文心蘭Oncidium ‘Gower Ramsey’的基因轉殖,將一段來自於阿拉伯芥第一條染色體上長度約550bp的DNA片段或另一段來自水母的綠螢光蛋白基因片段放入文心蘭Oncidium‘ Gower Ramsey’中作為此品系的分子標誌。將PLB藉由農桿菌轉殖後以Hygromycin作初步篩選,將存活的PLB培養成植株後再萃取其DNA進行PCR及南方墨點分析證明轉殖的片段確定有插入植株中。再將確定的轉殖株作葉片培養或芽接芽等方式,培育出其子代,再萃取其子代的DNA,以PCR及南方墨點分析確認轉殖的片段仍然有存在於其子代,以證明此片段是可以穩定的遺傳到其增殖的子代。
Orchids are primarily grown for their large, long-lasting and fascinating flowers; thus, the improvement of quality attributes such as flower color, longevity, shape, architecture, biotic and abiotic stress tolerance and the creation of novel variation are important economic goals for floriculturists around the world. However, until now much attention has not focused on protection of novel cultivars from the unauthorized breeders to safeguard our orchid industry economy. Recent advances in genetic engineering and molecular biology techniques augmented with gene transformation could help growers to protect novel cultivars in the competitive orchid industry. Henceforth, we initiated a systematic study to develop novel method to protect the identity of commercially important orchid cultivar by the introduction of foreign DNA sequence as molecular marker via Agrobacterium-mediated transformation method in Oncidium Gower Ramsey and optimization of transformation conditions. A unique untranslated Arabidopsis DNA sequence (550bp) or a humanized Renilla green fluorescent protein fragment without promoter was transformed into Oncidium (Gower Ramsey) by Agrobacterium-mediated method. Around 60 independent transgenic lines were generated, integration of molecular marker (550bp) in the genome of some transgenic orchids were confirmed by PCR analysis with specific primers. Taken together, the developed technology system could be successfully employed in protecting rights of Taiwan orchid grower’s in the international market from the unauthorized breeders.
中文摘要………………………………………………………………Ι
英文摘要………………………………………………………………Π.
目錄……………………………………………………………………Ⅲ
表目錄…………………………………………………………………Ⅴ
圖目錄…………………………………………………………………Ⅵ
附錄目錄………………………………………………………………Ⅷ


第一章 前言……………………………………………………..…1
第二章 文獻探討
2.1 文心蘭簡介…………………………………….…..…3
2.2 文心蘭的型態與生長習性…………………….……..3
2.3 文心蘭常見的品種…………………………….……..4
2.4 植物基因轉殖之研究………………………….……..4
2.5 品種鑑定………………………………………….…12
第三章 材料與方法
3.1 文心蘭材料…….………………………………....…15
3.2 農桿菌品系………….……………….………….…..15
3.3 農桿菌轉殖的方法……….……………………..…..15
3.4 基因轉殖植物抗生素篩選……..……………..….....16
3.5 轉殖株分析…………….....…………………..……..17
第四章 結果
Ι. Orchi550……………………….………………..……..23
Π. hrGFP…………………………………………..……..26
第五章 討論
5.1 農桿菌轉殖………………………………………….28
5.2 轉殖株分析…………………………………….……29
5.3 分子標誌的選擇…………………………………….30
5.4 PCR在品種鑑定上的重要性……………………...31
第六章 參考文獻…………………………………………………33
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