分析水稻T-DNA插入突變株: M0022150, M0023563, M0023580, M0037352及M0032079 Analysis of T-DNA insertion mutants:M0022150, M0023563, M0023580, M0037352 and M0032079

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Title:	分析水稻T-DNA插入突變株: M0022150, M0023563, M0023580, M0037352及M0032079 Analysis of T-DNA insertion mutants:M0022150, M0023563, M0023580, M0037352 and M0032079
Authors:	吳欽鋒;Chin-Feng Wu
Contributors:	生命科學研究所
Keywords:	T-DNA;水稻;rice;T-DNA
Date:	2009-01-16
Issue Date:	2009-09-22 10:21:01 (UTC+8)
Publisher:	國立中央大學圖書館
Abstract:	水稻是重要的糧食作物，並且是可以探討水稻基因對於植物生長及糧食生產的單子葉模式植物。隨著水稻基因體解碼計畫的完成，下一個挑戰便是大規模地探討基因的功能性。功能性基因體突變庫則提供了有效的研究基因功能的工具。在台灣的水稻突變體庫（TRIM）利用T-DNA插入性突變法產生了55,000株包含啟動子捕捉、基因活化、基因插入性缺失的突變體，因此是很好用來研究水稻功能性基因體的來源。在本論文研究裡，我們從突變體庫裡挑選5株在葉片外表型具有褐斑或是白化的突變株進行研究，利用質體救援可以找到T-DNA的插入點，並藉由南方墨點法來偵測T-DNA的插入數目。我們選取的突變株經由實驗結果發現T-DNA皆插在基因與基因間，並且分析突變株的後代子群體插入點及T-DNA插入點鄰近的基因表現量時，卻發現突變株所造成的褐斑或白化性狀，並非是因為T-DNA插入位置所造成的。在M0032079突變株分析上，發現Os05g36990被T-DNA上的加強子所活化，因而在葉子上有較高的表現量，而此基因為植物專一的ovate domain，並且在水稻莖、桿及穗有專一性表現，但其基因在植物體所扮演的功能卻是未知。因此嘗試以細胞定位及觀察過量表現OsOFP基因轉殖株之性狀，探討OsOFP可能之生理意義。在本論文中利用基因槍將OsOFP與GFP之融合基因轉殖於洋蔥表皮細胞，由GFP螢光位置以得到OsOFP在細胞中之定位，結果發現OsOFP在細胞核與細胞質中均有表現。另一方面，利用農桿菌轉殖法得到過量表現HA-OsOFP之阿拉伯芥T1轉殖株，發現此基因可能在植物生長調控細胞延長上扮演一個抑制角色。而這樣的結果可以幫助我們瞭解OsOFP在植物訊息傳遞路徑上扮演何種角色。未來更可以利用大量表現HA-OsOFP與基因靜默的轉殖珠相互比較，以幫助我們釐清OsOFP在植物體內的功能。 Rice, as an important crop and a model plant, provides major insights into gene functions for crop growth and production. With the completion of a rice genome sequencing project, the coming challenge is to determine gene functions via large scale analysis. Phenomics with detailed information about tagged populations provides a good tool for functional genomics analysis. Taiwan Rice Insertion Mutants (TRIM) has generated a rice mutant population, containing 55,000 promoter trap and gene activation or knockout lines by a T-DNA insertional mutagenesis approach, and offers a good resource for rice phenomics study. In this study, five T-DNA inserted mutants were selected with leaf-lesion-mimic or albino phenotypes. T-DNA insertion loci of these mutants were identified by plasmid rescue and have two or three T-DNA copy numbers by southern blot assay. We found that T-DNAs were inserted at inter-gene space among these selected rice mutants. Co-segregation analysis of progenies and RNA expression levels of genes near the T-DNA insertion locus were show that the lesion-mimic or albino phenotype of these mutants was not caused by T-DNA inserted loci. In M0032079, we found that Os05g36990 was highly expressed in its leaves. Os05g36990 encode a plant specific ovate domain and expressed in stems, stalks and panicles. However, its function is unclear. In this study, we tried to find the functions of OsOFP by the tissue or subcellular distribution of GFP-fusion protein expressed in transgenic plants followed by phenotype analysis. Subcellular localization of OsOFP was determined by particle bombardment, our results showed that OsOFP-GFP fusion protein localized at the nucleus and cytoplasm in onion epidermal cells. We established a transgenic T1 plant overexpressing HA-OsOFP by Agrobacterium-mediated transformation. Our results indicated that OsOFP may function as a repressor in regulating cell elongation in plants. These results can help us to understand the possible roles of OsOFP in the signal transduction. The overexpressing HA-OsOFP transgenic T1 plants can compared with RNAi transformants in the future and help us to further identify its function.
Appears in Collections:	[Graduate Institute of Life Science] Electronic Thesis & Dissertation

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