稻属钙调素基因家族之分子演化研究.DOC

稻屬鈣調素基因家族之分子演化研究 Molecular Evolution of Calmodulin Gene Family in Oryza L. 研究生：林麗文 Lin, Li-Wen 指導教授：江友中 Chiang, Yu-Chung 【摘要】 稻米是世界上最主要的糧食作物之一，全球有二十億以上的人口以稻米為主食，因此廣被學者研究。目前已有研究發現，細胞外不同的刺激和鈣離子/鈣調素(calmodulin)有密切的關係，鈣離子是生物體內重要的二級傳導物，其中鈣調素是真核生物中最佔優勢的接受器，參與植物多種生理上調節，包含乾旱、淹水、高鹽、和植物生長調節劑等環境逆境之訊息傳導。但是在分子演化方面並不清楚，本研究主要針對稻屬植物、高山旱稻、栽培稻、野生稻等模式植物去探討，我使用聚合酶連鎖反應和選殖定序技術獲得三個鈣調素基因座，分別位在第三、五、十二號染色體，並進行親緣關係分析(phylogenetic analysis)、遺傳歧異度分析(genetic diversity)、中性假說測定(neutrality test)及最大相似度親緣分析(phylogenetic analysis by maximum likelihood, PAML)。遺傳歧異度在染色體十二號各類群分析中，栽培稻、當地稻和旱稻類群的數值較野生稻為低，推測因為人類馴化導致遺傳變異降低。在中性假說測試中，第十二號染色體中皆符合中性；第三號染色體在所有稻屬中其全長及內含子(intron)些微偏離中性；在第五號染色體所有稻屬中其各個區域皆偏離中性。由第五號染色體推論為單一點突變過高所造成負值，依據中性假說中外顯子與內含子機率應相同，表示每一世代所累積的突變，因為純化天擇，導致外顯子中之非同義取代被移除，同義性取代乃存在，在內含子中突變持續累積，因此造成現今現象。在最大相似度分析中，三個基因座皆無正向天擇的現象。 關鍵字：鈣調素，稻屬，亞洲野生稻，旱稻，抗旱，分子演化 【Abstract】 Rice is the most important and staple crop in the world. More than two billion people eat rice as a major source of carbohydrate. In recent research, calmodulin (CaM) is an important protein for signal transduction and response to different stress including drought, flooding and saline soil in plant. In order to understand the regulation of stress resistant in plant, it is important to study on the molecular evolution in the gene family of calmodulin. The genus of Oryza, with two crop species and 21 wild species, is the famous genus in Poaceae. With the complete rice genome data, several genes of camodulin are identified using bioinformatics.In this study, we try to clone different calmodulin genes from different species in the genus Oryza, upland , cultivative and wild rice. We used polymerase chain reaction, cloning and sequencing techniques to obtain three calmodulin gene family which are located in Third, Fifth and the 12th chromosome respectively, then we take phylogenetic analysis, genetic diversity analysis, neutrality test and phylogenetic analysis by maximum likelihood (PAML). Compared to the genetic diversity of 12th chromoso