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National Chung Hsing University Institutional Repository - NCHUIR > 農業暨自然資源學院 > 生物科技學研究所 > 依資料類型分類 > 碩博士論文 >  阿拉伯芥中調控花藥開裂與花粉發育之LEA40 基因之分子選殖與功能性分析

Please use this identifier to cite or link to this item: http://nchuir.lib.nchu.edu.tw/handle/309270000/152589

標題: 阿拉伯芥中調控花藥開裂與花粉發育之LEA40 基因之分子選殖與功能性分析
Characterization and functional analysis of LEA40 gene in controlling anther dehiscence and pollen development in Arabidopsis thaliana
作者: 黃凱郁
Huang, Kai-Yu
Contributors: 楊長賢
Chnag-Hsien Yang
生物科技學研究所
關鍵字: 阿拉伯芥;花藥;花粉
Arabidopsis thaliana;anther;pollen
日期: 2012
Issue Date: 2013-11-18 10:44:19 (UTC+8)
Publisher: 生物科技學研究所
摘要: LEA (late embryogenesis abundant)蛋白之功能被報導與植物發生失水及霜害的耐受性有關。LEA蛋白可以單獨的保護蛋白質在乾燥及冰凍的過程中保持構型。LEA40 是阿拉伯芥中LEA蛋白家族中的一員,透過real-time PCR 發現,LEA40基因的 mRNA在年輕的小花苞中有高量的表現。而根據公開的微陣列(microarray)資料(eFP browser) 得知LEA40基因在花粉具有專一的表現位置。為了探討LEA40基因表現位置與功能的相對應關係,本研究利用農桿菌媒介轉殖法產生基因轉殖植物以進行其功能分析。以35S啟動子大量表現LEA40的轉殖株,有著花藥不開裂造成雄不稔及開花時間提早之現象。進一步利用亞力山大(Alexander’s)染色法,發現其花粉活性外觀與野生株花粉無異。為了瞭解花藥不開裂的原因,使用茉莉酸(Jasmonic acid, JA) 處理後,花藥仍然不開裂。此結果排除了JA缺乏造成性狀的因素。相對的,利用RNA干擾產生的LEA40 基因表現下降的轉殖株,亦有雄不稔的現象。利用亞力山大染色及雷射掃描共軛焦顯微鏡觀察LEA40 RNAi的花粉顆粒,發現其有嚴重皺縮的缺陷出現。這個結果指出LEA40基因的功能可能是藉由在分子間穩定細胞的結構,進而保護蛋白質在脫水的過程能保持構型,間接調控花藥的開裂與花粉的成熟。
LEA (late embryogenesis abundant) proteins in plants are associated with tolerance to water stress resulting from desiccation and cold shock. LEA proteins can protect protein from aggregation due to desiccation and freezing. LEA40 is one of the members of LEA protein family in Arabidopsis.High level of LEA40 mRNA was found in young floral bud by real-time PCR analysis. According to public microarray data (eFP browser) the expression of LEA40 was specifically detected in pollen. To explore the relationship between function and expression pattern of LEA40 gene, Agrobacterium-mediated plant transformation in Arabidopsis was performed. Ectopic expression of LEA40 caused a male sterility phenotype with anther indehiscence and early flowering in trangenic Arabidopsis. Pollen grains of 35S::LEA40 transgenic lines were viable and did not show cytoplasmic defect by Alexander’s staining. Further analysis indicated that the indehiscence of the anther in 35S::LEA40 Arabidopsis was not caused by the alteration of JA signaling pathway. By contrast, RNAi mediated LEA40 gene depletion caused early dehiscence of anther and the defect of pollen, resulting in the male sterility in trangenic Arabidopsis plants. This result indicated that LEA40 protein may stabilize cellular structures and protect protein from aggregation due to the desiccation during anther dehiscence and pollen development.
Appears in Collections:[依資料類型分類] 碩博士論文

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