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研究生:林郁姍
論文名稱:白腹叢蚊原酚氧化酵素Ⅲ基因的選殖及特性的分析
論文名稱(外文):Cloning and characterization of Armigeres subalbatus prophenoloxidase Ⅲ gene
指導教授:陳正成陳正成引用關係
指導教授(外文):Cheng-Chen Chen
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:寄生蟲學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2003
畢業學年度:91
語文別:中文
論文頁數:58
中文關鍵詞:原酚氧化酵素黑化反應白腹叢蚊基因調控啟動子活性分析
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原酚氧化酵素為昆蟲黑化反應 (melanization)的關鍵酵素,在昆蟲體內參與許多重要的生理功能,例如:寄生蟲之黑化反應,傷口癒合,表皮骨骼化,卵殼硬化等。本實驗室以自吸血之白腹叢蚊(Armigeres subalbatus)中選殖一個原酚氧化酵素之cDNA,將之命名為As-pro-PO III,我們利用RT-PCR技術檢測As-pro-PO III在不同的生理狀況下,例如吸血、傷口癒合、感染細菌或微絲蟲均是維持著一個持續性非引導性的表現。為了研究As-pro-PO III之基因調控機制,我們利用基因庫篩選的方法,成功的選殖到As-pro-PO III基因,經過電腦分析其序列,發現As-pro-PO III含有四個內插子(長度分別是59,61,53,64 bp),利用引子延伸方法來分析As-pro-PO III的轉錄起始點在距離轉譯起始點上游-50處。我們將As-pro-PO III上游約1600bp序列進行啟動子活性分析,得到-279至-36可能為啟動子區域。經電腦分析此段序列,結果發現數個可能的調控子結合序列:GATA、AP-1、Dfd、zeste。

The production of melanotic material is an important physiological process in various aspects of insect life, i.e. cuticular tanning between moults, egg chorion tanning, wound healing, and melanotic encapsulation. One of the key enzyme involved in the production of melanin is phenoloxidase (PO). Recently, a prophenoloxidase cDNA(As-pro-PO III) was cloned from blood-fed mosquito Armigeres subalbatus. In this study, RT-PCR analysis demonstrates that As-pro-PO III were persistently expressed in mosqutos, but not up-regulated in many physiological conditions, i.e. blood-feeding, wound healing, microfilaria and bacteria inoculation. Subsequently, the gene of As-pro- PO III was obtained by genomic library screening. As-pro-PO III gene contains four introns that are 59,61,53,64 bp long. The transcription starting site was on the -50 upstream from the translation starting site. Cell transfection assay suggests that the region from -279 to -36 contains regulatory motifs. In this 5’ flanking sequence, we found several putative regulative motifs, i.e. GATA, AP-1, Dfd, zeste,etc.

中文摘要………………………………………………………………1
英文摘要………………………………………………………………2
第一章、緒論………………………………………………………3-7
第二章、研究動機……………………………………………………8
第三章、研究目的……………………………………………………8
第四章、研究流程……………………………………………………9
第五章、材料與方法………………………………………………10-25
第六章、結果………………………………………………………26-30
第七章、討論………………………………………………………31-34
第八章、參考文獻…………………………………………………35-39
第九章、圖表及說明………………………………………………40-54
第十章、附錄………………………………………………………55-58

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