豆類作物間之簡單重複序列及其相鄰區域之變異

Variation of Simple Sequence Repeated DNAs and their Flanking Regions among Legume Crops

楊欣佳(Sin-jia Yang)

指導教授 ： 林順福

國立臺灣大學/生物資源暨農學院/農藝學研究所/碩士(2005年)

https://doi.org/10.6342/NTU.2005.10337

摘要

本試驗目的在利用ISSR及SSR DNA分子標誌來探討：(1)豆類物種間簡單重複序列及其鄰近區域之DNA序列變異 (2) SSR及ISSR DNA分子標誌跨物種利用之可行性 (3)豆類物種間之分類或是演化關係；調查材料包括7個屬12個物種，即紅豆、綠豆、大豆、花生、菜豆、紅花菜豆、豇豆、豌豆、蠶豆、白鳳豆、紅鳳豆及樹豆等作物。利用SSR分子標誌轉移利用率評估引子設計之來源物種和其他物種間的相似程度，發現豌豆和大豆基因組間在引子黏合序列上保守性較高。結果證實SSR分子標誌在跨物種轉移利用是可行的，本試驗亦篩選得到兩組可在12個物種以及禾本科水稻共同利用之SSR引子。也發現在不同物種所增幅長度相近的專一性片段間，亦存在多量的DNA序列變異，而僅利用大豆AG81引子所增幅之DNA序列變異資訊，即可區分遠緣物種。由於豆類物種間的基因組結構差異甚大，因此本試驗篩選得到大量的物種特有之ISSR DNA分子條帶，可利用於物種間或是屬間物種的遺傳變異分析。而在遺傳相似度、群集分析或主成分分析結果均顯示豇豆屬內的紅豆、綠豆及豇豆三個物種之遺傳距離甚遠，和屬間物種的遺傳距離相近，故試驗結果傾向支持應將三個物種進一步區分之建議。SSR DNA序列分析以及ISSR分析的結果均顯示蠶豆與其他調查之豆類作物間有最大之遺傳距離，在蠶豆品種之AG81 DNA片段中，簡單重複序列結構幾乎已經不存在，亦可說明蠶豆較其他物種在簡單重複序列區域具有較大之DNA序列變異。本試驗結果不但證實ISSR分子標誌可有效利用於分析豆類物種間之遺傳變異，亦顯示SSR DNA分子標誌甚具有跨物種應用的潛力。

關鍵字

豆類作物 ； 簡單重複序列

並列摘要

To study genetic variation among legumes, ISSR and SSR DNA markers were analyzed around simple sequence repeats and their flanking regions for (1) the variation of simple sequence repeated DNAs and their flanking regions among legume crops (2) the feasibility of applying SSR and ISSR DNA markers in different species of legumes (3) the phylogenetic analysis of legume crops. Twelve important legume crops (33 varieties), including soybean (Glycine max), peanut (Arachis hypogaea), small red bean (Vigna angularis), mungbean (V. radiata), cowpea (V. unguiculata), pea (Pisum sativum), broad bean (Vicia faba), jack bean (Canavalia ensiformis/ gladiata), common bean (Phaseolus vulgaris/ coccineus) and pigeon pea (Cajanus cajan), were investigated. The transferability of SSR DNA markers was also used to evaluate the genetic similarity between the core species (soybean), which the SSR primesrs were originally designed, and other species. The conservative in primer annealing regions was found in pea and soybean. The experimental results indicate that it is applicable of using SSR markers of a species for the other legumes. In this study, two primer pairs able to amplify SSR sequences for 12 legume crops and rice were discovered. Also, lots of DNA sequence variations were identified among the similar length amplicons of the legume crops. And the distant species could be by discriminated by merely using DNA sequences amplified by soybean AG81 primer pairs. Because of the divergence in size of legume genomes, a great deal of ISSR primers were identified for a specific crop. These primers are available for detecting genetic variation among or within legume crops at molecular level. Because the long genetic distance among small red bean, mungbean and cowpea was found in the genetic similarity, cluster analysis and principle component coordinate analysis, each of the three species was suggested to become an isolated genera. According to SSR DNA and ISSR analysis, the broad bean had the largest genetic distance to the other legume crops. The finding of the degenerated repeated sequence in the AG81 segments of the broad bean also indicated that lots of DNA sequence variation had occurred in the region. Results from this study not only have proved the usefulness of ISSR DNA markers in detecting genetic variation among legume crops, but also have demonstrated the potential of applying SSR DNA markers in the other species.

並列關鍵字

Legume Crops ； SSR ； ISSR

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