苦瓜性別表現相關蛋白質之研究

Studies on sex expression-related proteins in bitter gourd (Momordica charantia)

賈儒珍(Ju-Chen Chia)

指導教授 ： 黃鵬林 ； 杜宜殷

國立臺灣大學/生物資源暨農學院/園藝學研究所/碩士(2005年)

https://doi.org/10.6342/NTU.2005.10394

摘要

苦瓜（Momordica charantia L.）為雌雄異花同株之葫蘆科作物，施用植物生長調節劑可調節雄雌花比例，但其機制目前尚無定論，本論文以蛋白質二維電泳為分析方法，獲得苦瓜性別表現相關蛋白質。調查月華品種苦瓜的花芽外型、長度，並以埋蠟切片觀察花芽發育狀況，歸納出雄花和雌花的四個發育階段，作為分析不同發育階段蛋白質表現之標準。以根、莖、葉、果實、雄花和雌花之蛋白質萃取物進行二維電泳及比對工作，歸納出8個花器相關蛋白質、15個雄花相關蛋白質、和12個雌花相關蛋白質，並依序編號。雄花不同發育階段的二維電泳結果可比對出51個具有差異性表現的蛋白質，其中13個已歸類為雄花相關蛋白質、4個已歸類為雌花相關蛋白質。另外在雌花不同發育階段的二維電泳結果中，找出31個差異性表現的蛋白質，有5個屬於雌花相關蛋白質，一個屬於雄花相關蛋白質。進一步以0.54 mM naphthalene acetic acid（NAA）和0.12 mM激勃素（gibberellic acid, GA3）噴施於苦瓜8-10片葉大之幼苗，取頂芽進行二維電泳，得知NAA對數個花器相關蛋白質和雌性相關蛋白質有抑制作用，GA則有誘導作用。經整理各試驗得到性別表現相關蛋白質之分子量和等電點，並比較其在不同器官中的表現，可區分為四群：第一群蛋白質在花器發育四個階段中累積量相等；第二群蛋白質只在其中一個性別的發育過程中累積量變化；第三群蛋白質在雄花和雌花發育過程中同時有增加或減少的趨勢；第四群蛋白質在雄花或雌花發育過程中累積量上升，在另一性別發育過程中下降。前三群蛋白質可能與雄花或雌花之生長發育相關，但與花性變化之關聯性較低；第四群蛋白質在兩性器官中有相反的表現，可能參與花性調控之作用機制。由以上結果得知，與花芽生長發育相關之蛋白質中，與花性變化密切相關者為18號、70號及74號蛋白質。日後可針對此蛋白質進行胺基酸定序，設計寡核苷酸探針，進行性別相關基因選殖工作。

關鍵字

苦瓜 ； 性別 ； 蛋白質

並列摘要

Sex expression of bitter gourd (Momordica charantia L.), a monoecious species in Cucurbitaceae, is altered by exogenous application of plant growth regulators, with unkown mechanism of sex determination. In this study, we use 2-dimensional protein gel electrophoresis as the analysis method of sex expression-related protein in bitter gourd. To conclude different development stages in male and female flowers for analysis of protein expression, the morphology, length and microscopic structures of flower buds in M. charantia cv. Moon Shine were observed. Roots, stem, leaves, male and female flowers were used as materials for 2-dimensional electrophoresis. Among these, 8 flower organ-related proteins, 15 male flower-related proteins and 12 female flower-related proteins were obtained and numbered. In different development stages of male flowers, 51 development-distinctive proteins were found; 13 of these are male flower-related and 4 are female flower-related proteins. During female flower development, 31 development-distinctive proteins were found; 5 of these are female flower-related proteins, and 1 are male expression-related protein. On the other hand, shoot apex of bitter gourd at the 8-leaf stage treated with 0.54 mM naphthalene acetic acid (NAA) and 0.12 mM gibberellic acid (GA3) were used to analysis for protein expression. NAA inhibited some of the flower organ-related proteins and female flower-related proteins, and GA could induce them. Based on the molecular weight, pI value and expression level of the proteins, four expression type of sex expression-related proteins were defined. Group 1 proteins expressed constitutively during flower development. Protein amount of group 2 changed during male or female flower development. Group 3 proteins elevated during male and female flower development. Group 4 proteins elevated during female flower development and declined during male flower development. Group 1~3 proteins were possibly involved in male or female flower development, but less related with sex-differentiation. Group 4 proteins were found highly related to sex-determination due to their expression characteristics. The results showed that spot 18, 70 and 74 had close relationship with sex-determination and could be used for cloning of sex expression-related genes.

並列關鍵字

bitter gourd ； sex ； proteins

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