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研究生:陳哲仁
論文名稱:黃肉蕃茄黃色素基因座之遺傳分析
論文名稱(外文):Candidate gene analysis of yellow pigmentation loci in yellow flesh tomato
指導教授:古新梅
學位類別:碩士
校院名稱:國立中興大學
系所名稱:農藝學系
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:2003
畢業學年度:91
語文別:中文
論文頁數:66
中文關鍵詞:番茄黃肉基因座增殖片段長度多型性
外文關鍵詞:AFLPSCAR
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類胡蘿蔔素是高等植物重要的化學成分,類胡蘿蔔素結構基因的表現,和類胡蘿蔔素組成與成份比例,影響成熟番茄果實的顏色,本實驗以增殖片段長度多型性(簡稱AFLP)分析不同顏色的番茄品種,包括本地番茄亞蔬六號和自田間發現的黃肉種YM品系,與日本商用黃色番茄F1 yellow pico(簡稱YP),以及八個種源中心的果色突變體,利用遺傳分析方法找尋與YM品系黃肉性狀相關的分子標誌,本實驗一共使用62個不同的AFLP引子組合,結果不同品系約測得1100個條帶,平均一個AFLP引子組合測得18個左右的條帶,並以亞蔬六號測得條帶作為對照,計算其他品系的多型性條帶,以本地的YM品系測得45個多型性條帶為最少,約佔整體4%,至於其他參試品系則超過16%,進一步以YM x YP F2雜交子代作為分析族群,根據子代果實顏色分為紅和黃色兩群,各逢機挑選7個個體混合成子集團,進行AFLP分析,發現6個AFLP分子標誌與黃肉性狀有關,將這些與果肉顏色性狀有關的AFLP分子標誌作選殖及解序,並未直接發現與任何已知類胡蘿蔔素生合成基因有關,以及這些分子標誌在番茄染色體的位置,接著將這六個AFLP分子標誌轉換成分析上較簡便的sequence-characterized amplified region(簡稱SCAR)標誌,結果發現分子標誌SCAR-E8M1與黃肉性狀具有連鎖關係,可作為黃肉基因座據圖選殖的基礎,和分子標誌輔助育種的利用。
Caroteniods are essential compounds of higher plants. Expression of caroteniods biosynthesis genes, compose of caroteniods and the ratio inferences tomato fruit color. In this study, we used amplified fragment length polymorphism (AFLP) assay different tomato varieties including local AVRDC No. 6 and YM, commercial yellow hybrid called yellow pico (YP) ,and in addition, eight tomato color mutants. Genetics analysis was performed to survey yellow flesh trait associate markers, a total of 62 different AFLP primer combinations was used. About 1100 bands were detected among these varieties and on average, 18 fragments were amplified per primer combination. Based on pattern of AVRDC No. 6, we detected less than 4% polymorphic bands in local tomato YM variety and over 16% polymorphic bands in other tested varieties. We also surveyed F2 population derived from YM and YP. According to fruit color of progenies, 7 individuals were randomly mixed as bulk. YM, YP, and two bulks were tested and detected 6 polymorphic fragments. After the fragments were subcloned and sequenced , no any significant similarity between carotenoids synthase genes or the location on the tomato chromosome were found. The polymorphic fragments were transferred into SACR markers. One marker SCAR-E8M1, was found associate to yellow fresh trait. The markers obtained in this study could be used in map-based cloning and marker-assisted breeding.
中文摘要_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ v
英文摘要_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _vii
壹、前言 _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 1
貳、前人研究 _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 3
一、類胡蘿蔔素之重要性_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _3
二、類胡蘿蔔素生合成途徑之研究_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 3
三、番茄果實成熟期間類胡蘿蔔素含量之變化_ _ _ _ _ _ _ _ _ _ 6
四、類胡蘿蔔素突變體之研究_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _6
五、據圖選殖之研究_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _9
六、增幅片段長度多型性之研究_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _10
七、Bulked segregant analysis分析之研究_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 13
參、試驗材料與方法_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 16
一、植物材料_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 16
二、總量DNA之萃取_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 16
三、增幅片段長度多型性分析AFLP_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 17
四、聚丙醯胺膠體電泳和銀染法_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 18
五、聚丙醯胺膠體上差異片段之回收_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 19
六、差異片段之擴增及選殖_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 20
七、少量質體DNA之抽取_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 21
八、殖入片段之篩選及核酸自動定序分析_ _ _ _ _ _ _ _ _ _ _ _ 22
九、顏色相關殖系之序列分析_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 23
十、Sequence-Characterized Amplified Region分析_ _ _ _ _ _ _ 23
肆、結果_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 25
一、YM果實外表型及黃色基因座分析_ _ _ _ _ _ _ _ _ _ _ _ _ _ 25
二、番茄不同果實顏色品種(系)AFLP分析_ _ _ _ _ _ _ _ _ _ _ 25
三、YM x YP F2子代AFLP / BSA分析_ _ _ _ _ _ _ _ _ _ _ _ _ _ 26
四、黃肉性狀相關殖系之序列分析_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 27
五、黃肉性狀相關片段之SCAR分析_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 29
伍、討論_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 32
一、YM品系非r基因座突變體_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 32
二、番茄不同果品種(系)遺傳組成之差異_ _ _ _ _ _ _ _ _ _ _ 32
三、番茄不同果品種(系)AFLP分析之差異_ _ _ _ _ _ _ _ _ _ _ 33
四、YM品系可能與黃肉性狀相關之AFLP分子標誌_ _ _ _ _ _ 34
五、黃肉性狀相關AFLP分子標誌序列分析_ _ _ _ _ _ _ _ _ _ _ 35
六、SCAR分子標誌選拔性狀_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 37
七、參考文獻 _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ 40
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