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研究生:潘慶芝
研究生(外文):Chin-Chih Pan
論文名稱:台灣落花生品種分子鑑定之研究
論文名稱(外文):Studies on Molecular Identification of Taiwan Peanut Varieties
指導教授:陳成陳成引用關係林順福
指導教授(外文):Cheng ChenShun-Fu Lin
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:農藝學研究所
學門:農業科學學門
學類:一般農業學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:79
中文關鍵詞:落花生分子鑑定品種鑑別
外文關鍵詞:peanutmolecular identificaitonvarieties idetification
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為防止落花生走私進口與確保國產落花生之遺傳特性及品質,本試驗進行利用分子標誌鑑別國產落花生品種之研究。本研究以國內17個主要之落花生栽培品種為材料,探討利用不同處理方法由種仁抽取DNA之可行性,並利用分析方便、穩定性高且易於推廣之ISSR分子標誌進行落花生品種指紋資料庫建立,及遺傳相似性分析並建立品種之鑑定流程;另以三個栽培種及其親本為材料,探討以雜交親本分子特性供品種鑑定之可行性。此外,本研究亦探討利用cDNA進行遺傳相似性分析結果與利用ISSR DNA逢機偵測所得遺傳相似性結果之異同,以期瞭解此法在指紋分析與品種鑑定之發展潛力。主要研究結果顯示如下:
1. 不同處理抽取種仁DNA所得結果在訊號較強之引子皆相同,但在訊號較弱之引子則有優劣之分,本研究結果推薦由胚直接抽取DNA,可快速得品質佳之DNA。
2. ISSR分子標誌可有效將國內主要栽培品種區分出,並可得具品種專一性之分子標誌,據此利用6個引子產生之12個分子標誌可建立一套有效品種鑑定流程。
3. 利用ISSR分子標誌所得落花生栽培品種遺傳變異介於5 % ~ 58 %,顯示國內栽培種落花生利用引種與雜交育種已逐漸擴充遺傳變異範圍。
4. 由雜交親本分子特性可得知育成品種之親本貢獻率及選拔偏差,亦可輔助品種鑑別,但需注意約有0.94 % ~ 4.02 %之異於親本條帶產生。
5. 偵測cDNA片段所得遺傳相似性結果與ISSR相符,但因cDNA手續繁複與費用高昂之缺點,較適宜針對特定功能之基因分析。
To prevent smuggling import and to ensure the genetic characteristic, molecular markers were used to identify the peanut varieties cultivated in Taiwan. Methods for extracting DNA from seeds were predetermined. Since the convenience in analysis, high stability and easy to extension of ISSR (inter simple sequence repeat) markers, this system was employed to establish DNA fingerprinting, to analyze genetic similarity, and to propose a feasible scheme for variety identification. In addition, 3 cultivars and their parents were used to study the possibility of identifying varieties with marker profile of their parents. Results of genetic similarity analysis by ISSR markers was also compared with that of cDNA ones to evaluate the potential of cDNA markers applied in variety identification. The results of this study were summarized as following:
1. The markers with strong bands were detected in different treatments for DNA isolation from peanut seeds, but for the weak bands were only detected in 3 of the 7 treatments. With better quality of PCR products, the fast method of DNA isolation from embryo of seed was recommended from this study.
2. The peanut varieties cultivated in Taiwan were identified with ISSR markers, and specific markers for varieties were found. An efficient scheme for variety identification was recommended by the use of 6 ISSR primers producing 12 informative markers in total.
3. Genetic distance among investigated varieties was detected from 5% to 58% on ISSR marker basis. It indicate that the genetic variation of varieties cultivated in Taiwan have became more diversified through continuous plant introduction and crossing breeding.
4. Selection declination was discovered from the comparison of ISSR markers between varieties and their parents. About 0.94%~4.02% markers of 3 varieties were not inherited from both of their parents implying possible error of identifying varieties from ISSR markers of parents.
5. Similar result of genetic similarity study was found between the cDNA and ISSR markers. Due to the complicated process for operation and more expensive chemicals used, the cDNA markers are suitable for analyzing specific genes rather than detecting genetic diversity.
一、前言‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 1
二、前人研究‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 4
三、材料與方法‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧11
(一)落花生種仁抽取技術之探討‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧11
(二)落花生品種ISSR DNA指紋資料庫之建立及遺傳相似性分析
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧13
(三)落花生品種之ISSR分子標誌鑑定流程之建立
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧20
(四)利用cDNA分子標誌進行落花生品種遺傳相似性分析
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧20
(五)以雜交親本分子特性輔助品種鑑定
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧29
四、結果與討論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧32
(一)落花生種仁抽取技術之探討‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧32
(二)落花生品種ISSR DNA指紋資料庫之建立及遺傳相似性分析
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧36
(三)落花生品種之ISSR分子標誌鑑定流程之建立
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧46
(四)利用cDNA分子標誌進行落花生品種遺傳相似性分析
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧52
(五)以雜交親本分子特性輔助品種鑑定
‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧ 63
五、結論‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧69
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英文摘要‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧‧72
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