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研究生:白宗晏
研究生(外文):Zoung-Yan Bai
論文名稱:利用分子輔助選拔改善臺稉九號白葉枯病之抗性
論文名稱(外文):Improving bacterial blight resistance in TK9 rice variety by using marker-assisted selection
指導教授:王強生
口試委員:鍾美珠楊嘉凌林大鈞
口試日期:2016-07-29
學位類別:碩士
校院名稱:國立中興大學
系所名稱:農藝學系所
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:2016
畢業學年度:104
語文別:中文
論文頁數:103
中文關鍵詞:水稻育種白葉枯病米質
外文關鍵詞:RiceBreedingBacterial blight diseaseGrain quality
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水稻白葉枯病(bacterial blight disease, BBD) 為一種系統性病害,由白葉枯病菌 Xanthomonas oryzae pv. oryzae (Xoo)感染造成,病原菌經由傷口進入,移動至維管束繁殖並擴散,危害葉片、葉鞘與稻穗,為重大的水稻病害之一,嚴重危害水稻產量與米質。可經由田間管理、生物製劑、化學藥劑等方法防治,但成效有限且對於環境不友善。臺稉九號(TK9)是臺灣重要的良質米品種,具有良好株型、優良米質等農藝特性,但對白葉枯病抗性差,亟待改良。本研究以國際稻米研究所(International Rice Research Institute, IRRI)育成具有五個抗白葉枯病基因 (Xa4+xa5+Xa7+xa13+Xa21)的秈稻品種IRBB66作為抗病基因的提供親,與TK9進行雜交產生F1,連續與TK9回交至BC5世代,並於回交過程利用分子標誌輔助選拔 (marker- assisted selection,MAS)進行前景、米質及背景篩選,挑選出具有五個抗性基因之植株,使其自交純化,希望篩選具有抗病能力與TK9優良米質特性之植株,育成抗病之良質米品種。本試驗於2015第一期作,選育出18株具有三個、四個及五個同質結合抗性基因之 BC5F3植株,皆具有TK9良好米質基因型。於苗栗區農業改良場、霧峰農業試驗所、國立中興大學北溝農場、臺中改良場及臺南改良場多地區種植,並接種多株白葉枯病生理小種,抗病性檢定結果皆呈抗性反應,各地區抗病品系產量表現與TK9無差異。米質分析與食味品評結果,亦具有 TK9良好米質特性,未來完成可區別性(distinctness)、一致性(uniformity)及穩定性(stability)等DUS檢定後,即可申請水稻品種權,可成為抗白葉枯病新品種,或作為良好的雜交材料。

Bacterial blight disease (BBD) is a systemic disease which invaded the vascular tissue by Xanthomonas oryzae pv. oryzae (Xoo) and causes significant yield loss in rice production. However, it is ineffective and environmentally-unfriendly to control this disease by field management, antibiotics and chemicals. TK9 is one of the most popular japonica type rice varieties in Taiwan with good plant type, stable yield and excellent grain quality, but it is susceptible to bacterial blight disease. In this study, an indica-type rice variety, IRBB66, which harbors five BB resistant genes Xa4, xa5, Xa7, xa13 and Xa21, developed by International Rice Research Institute (IRRI) was used as a resistant donor parent. Crossing between TK9 and IRBB66 to was conducted generate F1 plant with five heterozygous resistant genes and followed by conventional backcrossing up to BC5F1 generation. Foreground, grain quality and background selections by the marker-assisted selection were applied to select the desired individuals with five resistant genes and high recovery percentage of recurrent parent, TK9. Eighteen BC5F3 resistant lines that posses three, four, and five homozygous Xa genes, and same genotypes of grain quality as TK9. These lines were planted in multi-location and consistently showed resistance against BB isolates. Their eating quality is similar with TK9, and can be developed as a high quality rice variety. Once complete with evaluation for distinctness, uniformity and stability, the DUS test, these TK9 bacterial blight resistance lines have the potential to become high quality varieties as well as hybrid parents.

壹、前言 1
貳、前人研究 2
一、水稻白葉枯病 2
二、水稻白葉枯病免疫反應之研究 3
三、水稻白葉枯病抗性基因之研究 5
(一) Xa4基因定位與抗性機制之研究 5
(二) xa5基因定位與抗性機制之研究 6
(三) Xa7基因定位與抗性機制之研究 7
(四) xa13基因定位與抗性機制之研究 8
(五) Xa21基因定位與抗性機制之研究 8
四、稻米品質 10
五、分子標誌輔助育種 13
六、基因堆疊 14
參、材料方法 17
一、試驗材料 17
二、試驗方法 18
1.水稻育苗及種植 18
2.育種流程 18
3.水稻基因組DNA之萃取 19
4.水稻小量基因組DNA之萃取 20
5.聚合酶連鎖反應(polymerase chain reaction, PCR) 21
6.聚丙烯醯胺膠體製作 22
7.聚丙烯醯胺膠體分析 22
8.洋菜膠體之製作 22
9.水平式電泳膠體分析 23
10.膠體染色與拍照 23
11.電泳資料分析 23
12.連鎖圖譜繪製 23
13.輪迴親回復率之估算 24
14.白葉枯病抗性檢定 24
15.菌流測試 25
16.農藝性狀調查 25
17.米質檢測 26
18.穀粒掃描與穀粒長、寬之測量 27
19.區域性試驗 27
20.統計分析 28
肆、結果 29
一、抗病品系之選拔 29
二、抗病性檢定 31
(一)參試品種 32
(二)TK9抗病品系 32
三、農藝性狀調查 33
四、米質分析 35
五、地區性試驗 37
(一)抗病品系於各地區之抗病表現 37
(二)抗病品系於各地區之產量表現 38
伍、討論 40
一、分子標誌輔助選拔 40
二、接種白葉枯病原菌之抗感分析 42
三、農藝性狀 43
四、米質分析 45
陸、結論 46
柒、參考文獻 47


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