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研究生:蔡皇龍
研究生(外文):TSAY, HWANG-LONG
論文名稱:阿拉伯芥中酵母菌SAC1同源性基因之分析
論文名稱(外文):Characterization of an Arabidopsis gene with homology to yeast SAC1 gene
指導教授:王淑美王淑美引用關係
指導教授(外文):WANG, SHUE-MEI
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:植物學研究所
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2001
畢業學年度:89
語文別:中文
中文關鍵詞:阿拉伯芥酵母菌SAC1
外文關鍵詞:ArabidopsisyeastSAC1
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摘要
阿拉伯芥多澱粉突變株sex4,將突變基因定位於第三號染色體m457、g4125標誌區中,在此標誌區中發現有一AccI限制脢切位在野生型及突變體中出現多型性的變異,而此變異落在SAC1-like基因座上。本論文分別選殖包含完整編碼區域不包含5’-或3’-端非轉譯區之SAC1-like基因genomic DNA片段,轉殖於突變株,在20株轉殖株中並未發現回復野生型之植株。另一方面利用RNAi的方式擬阻斷正常植株中SAC1-like基因的表現,觀察SAC1-like基因突變後的表現型,在73株轉殖株中未能觀察到外表型發生變化之突變株。由阿拉伯芥cDNA基因庫中選殖出SAC1-like基因的cDNA clone,並轉接於表現載體pGEX3KS大量表現蛋白質,並製備抗體,以SAC1-like抗體偵測SAC1-like基因在突變株中表現量較野生型低,並觀察植株中有同源蛋白質,分析SAC1-like蛋白質氨基酸序列,並於DataBase中發現,阿拉伯芥中還有K2A18.9與AtG5與SAC1-like有極高的相似性,且皆具有SAC1-like domain。由於SAC1-like基因在阿拉伯芥中之功能並不十分清楚,但在酵母菌中SAC1基因則參與訊息傳遞,將阿拉伯芥之SAC1-like基因之cDNA轉化至ΔSAC1缺失之酵母菌體中,結果阿拉伯芥之SAC1-like基因可以補償酵母菌SAC1基因突變之缺失,暗示阿拉伯芥之SAC1-like基因在植物中扮演類似SAC1的角色。

目錄
摘要………………………………………………………………
英文摘要………………………………………………………….
前言……………………………………………………………....
一、澱粉的代謝………………………………………………..
二、澱粉代謝之相關調控………………………………………
三、酵母菌SAC1與阿拉伯芥SAC1-like基因…………………..
材料與方法………………………………………………………….
一、阿拉伯芥DNA萃取………………………………………………
二、阿拉伯芥RNA萃取………………………………………………
三、北方墨點轉印分析法………………………………………………
四、南方墨點轉印分析法………………………………………………
五、放射線探針製備……………………………………………………
六、探針雜合反應……………………………………………………
七、阿拉伯芥cDNA 庫 SAC1-like基因之篩選……………………
八、不連續聚丙醯胺膠體泳動分析……………………………………
九、西方墨點轉印分析法………………………………………………
十、E. coli 質體DNA萃取………………………………………………
十一、SAC1-like基因抗體製備……………………………………
十二、酵母菌DNA轉化………………………………………………
結果………………………………………………………………
一、阿拉伯芥sex4突變株之特性…………………………….
二、SAC1-like基因cDNA之分離…………………………….
三、阿拉伯芥SAC1-like基因表現的情形……………………….
四、酵母菌SAC1基因缺失互補系統之建立…………………..
五、水稻基因組中亦有SAC1-like基因……………………….
附圖………………………………………………………………
討論………………………………………………………………
參考文獻………………………………………………………….

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