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標題: | 豆類Adh基因之DNA序列變異研究 Study on DNA Sequence Variations in Adh gene of legume species |
作者: | Yi-Hang Tseng 曾一航 |
指導教授: | 林順福 |
關鍵字: | 豆類,酒精脫氫酵素, legume species,alcohol dehydrogenase, |
出版年 : | 2007 |
學位: | 碩士 |
摘要: | 本研究目的在利用豌豆、阿拉伯芥、矮牽牛及草莓等四個雙子葉物種已知之Adh基因保守序列以選殖及定序紅豆之Adh基因,並且探討豆類物種Adh基因之DNA序列變異情形。試驗材料包括紅豆(8個品種)、綠豆(2個品種)、豇豆(2個品種)、菜豆(2個品種)、落花生(2個品種)、毛豆(2個品種)、蠶豆(2個品種)、紅鳳豆(1個品種)、白鳳豆(1個品種)及樹豆(1個品種),共計10個物種。利用兩組引子對完成紅豆(高雄5號)Adh基因之部分解序,其範圍包括部分intron 3至intron 9之間的區域,也印證利用基因之已知保守序列可選殖另一物種之相同基因,紅豆與豌豆之Adh基因序列比對結果顯示兩者之exon均較intron有較高之GC百分比(分別為41.0 ~ 48.7%及23.3 ~ 35.2%),且兩者在exon區域之DNA序列相似程度約80 %,而在intron區域中則有明顯的序列差異存在(長度差異最高可達116 bp),但是intron外側序列均具有GT/AG組成且相對較中央序列具有保守性。此外,比對豆類參試物種Adh基因之序列結果,發現Adh基因之intron 4與intron 6區域分別具有12及15 bp的InDel序列,且該intron區域在物種間或物種內皆有明顯的核苷酸變異。intron 4與 intron 6之序列雖然無法區分菜豆及豇豆,但可將紅豆、綠豆及其他物種區分為三個類群,故可利用於物種間之鑑別。而intron 4區域較適合於綠豆品種鑑定之用,及intron 6區域則適於菜豆品種之鑑別。由本研究結果可知,Adh基因之序列變異性具有分析物種間或物種內遺傳變異之應用潛力。 The purposes of this study were to clone and sequence Adh gene of azuki bean (Vigna angularis) according to the published sequences of the genes in four dicot species, including pea (Pisum sativum), arabidopsis (Arabidopsis thaliana), petunia (Petunia hybrida), and strawberry (Fragaria × Ananassa Duch.), and to scrutinize the sequence variation in legume species. A total of ten legume species were used as materials including Vigna angularis (8 varieties), Vigna radiate (2 varieties), Vigna unguiculata (2 varieties), Phaseolus vulgaris (2 varieties), Arachis hypogaea (2 varieties), Glycine max (2 varieties), Vicia faba (2 varieties), Canavalia gladiata (1 variety), Canavalia ensiformis (1 variety) and Cajanus cajan (1 variety). Segments of Adh gene in azuki bean (variety KS 5), covering an interval between partial intron 3 and intron 9, were amplified and sequenced with two pairs of primers. The result has proven the viability of using conserved sequences in a completely sequenced gene of one species to clone same gene in another species. In the comparison between Adh genes of azuki bean (Vigna angularis) and pea (Pisum sativum), high sequence similarity (about 80%) in exon and long length difference (up to 116 bp) in intron were found. Besides, the edge regions of Adh introns with GT/AG counterparts and more conserved sequences than central regions was observed in both species. In addition to two InDel (12 bp in intron 4, and 15 bp in intron 6), abound sequence variation in intron 4 and intron 6 of the investigated legume Adh genes. Sequence variations in these regions were able to discriminate azuki bean and mung bean (Vigna radiate), and other legumes but not difference between common bean (Phaseolus vulgaris) and cowpea (Vigna unguiculata). The intron 4 and intron 6 of Adh gene are respectively suitable for variety identification of mungbean and common bean. This study has demonstrated the application potential of Adh gene in genetic variation analysis among and/or within legume species. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/27988 |
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