Methionine sulfoxide reductase A (MSRA) 及 MSRB 修復 methionine-S-sulfoxide (Met-S-SO) 及 methionine-R-sulfoxide (Met-R-SO) 為 Met，單胞藻 (Chlamydomonas reinhardtii) 有 5 個 CrMSRA 及 4 個 CrMSRB isoforms。本研究探討高光調控 CrMSR mRNA 表現及光合電子傳遞鏈之角色? 研究過程 1、先設計可以區分不同 CrMSR isoform 之 quantitative real-time PCR primer，及設定 PCR 條件與產物序列確定後，進行 mRNA 分析。2、分析光合作用效率，光強 ≥ 300 μE m-2 s-1 及光合電子傳遞鏈抑制劑抑制光合氧氣釋放速率及 PSII 活性 (Fv/Fm, Fv´/Fm´)。3、基因表現量分析，單胞藻細胞由 50 μE m-2 s-1 轉移至 0、50、300、600 或 1,000 μE m-2 s-1，發現黑暗抑制 CrMSRA2, CrMSRA3, CrMSRB1.1, CrMSRB1.2, CrMSRB2.1 表現但誘導 CrMSRA4 表現，光強 ≥ 300 μE m-2 s-1 誘導 CrMSRA2, CrMSRA3, CrMSRA5, CrMSRB1.1,
CrMSRB2.1 及 CrMSRB2.2 表現而抑制 CrMSRA4 表現，與光強成正比，但不影響CrMSRA1 及 CrMSRB1.2 表現。外加 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) 抑制高光 (1,000 μE m-2 s-1) 效應。50 μE m-2 s-1 下，2,5-dibromo-3-methyl-6-isopropyl-p- benzoquinone (DBMIB) 促進 CrMSRA3、CrMSRA5、CrMSRB2.2 表現並抑制 CrMSRA4 mRNA表現。PSI electron donor，phenazine methosulfate (PMS，促進循環電子傳遞)，於 300 μE m-2 s-1 促進所有 CrMSRA isoform。所表現以，光合電子傳遞鏈 PQ 還原態 (-) 參與高光誘導 CrMSRA3、CrMSRA4、CrMSRA5、CrMSRB2.2 表現，Cyt b6f 還原態 (-) 參與 CrMSRA2、CrMSRB1.1及CrMSRB2.1。非循環電子傳遞鏈及循環電子傳遞鏈共同參與 CrMSR 基因表現調控。我們推測高光表現 CrMSR 與光氧化逆境性有關。

Methionine sulfoxide reductase A (MSRA) and MSRB are responsible for the repairing of methionine-R-sulfoxide (Met-S-SO) and methionine-S-sulfoxide (Met-R-SO) back to me-thionine, respectively. Five MSRA isoforms and four MSRB isoforms are discovered in the unicellular green alga Chlamydomonas reinhardtii. Whether high light regulates CrMSR ex-pression via photosynthetic electron transport (PET) was examined. By checking the se-quence of PCR product of each isoform, quantitative real-time primers were designed for discrimination of isoform expression. Light ≥ 300 μE m-2 s-1 and PET inhibitors inhibited PSII activity (Fv/Fm, Fv´/Fm´) and photosynthetic O2 evolution rate, particularly 1,000 μE m-2 s-1, in which it did not recover after 3 h. A transfer to dark decreased CrMSRA2, CrMSRA3, CrMSRB1.1, CrMSRB1.2, CrMSRB2.1 mRNA levels but increased CrMSRA4 mRNA levels. When exposed to 50, 300, 600, or 1,000 μE m-2 s-1, CrMSRA2, CrMSRA3, CrMSRA5, CrMSRB1.1, CrMSRB2.1 and CrMSRB2.2 mRNA levels increased as light ≥ 300 μE m-2 s-1, and concomitantly CrMSRA4 mRNA level decreased. Changes in mRNA levels increased as light intensity increased. The treatment of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) in 1,000 μE m-2 s-1 inhibited high light effect, and the treatment of 2,5-dibromo-3-methyl-6- isopropyl-p- benzoquinone (DBMIB) in 50 μE m-2 s-1 increased CrMSRA3, CrMSRA5 and CrMSRB2.2 mRNA levels but decreased CrMSRA4 mRNA level. The application of phena-zine methosulfate (PMS), an electron donor to P700+ that promotes cyclic electron transport, in 300 μE m-2 s-1 enhanced the increase of CrMSRA3 and CrMSRA5 mRNA levels by high light but inhibited the decrease of CrMSRA4 mRNA level, reflecting a role of cyclic PET. The above results let us to draw a conclusion that plastoquinone as reduced status mediates the expression of CrMSRA3, CrMSRA4, CrMSRA5 and CrMSRB2.2 by high light. The im-plication of linear electron transport and cyclic electron transport on the regulation of CrMSR gene expression will be discussed.We speculated that the high light up-regulation of CrMSR mRNA expression offers the resistance of Chlamydomonas to photooxidative stress.

目錄
頁次
口試委員會審定書
誌謝
中文摘要- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -- - I
英文摘要 (Abstract) - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -II
縮寫字對照- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - IV
目錄- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - VI
表目錄- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - VII
圖目錄- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -IX
附錄- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - XIII
壹、 前人研究- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -- - - - - - - - - - - - - -1
一、 光氧化逆境………………………………………………………………… 1
二、 Methionine (Met) 氧化與修補……………………………………………4
三、 Methionine sulfoxide reductase (MSR) 基因表現…………………… 7
四、 胞器與細胞核間基因表現之相互調控……………………………10
貳、 研究目的- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -- - - - - - - - - - - - - - 16
參、 實驗材料與方法- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -18
肆、 結果- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 29
伍、 討論- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - 52
陸、 參考文獻- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -59

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吳宗孟 (2009)。大型海藻裂片石蓴 (Ulva fasciata Delile) 過量銅毒害之基因表現。國立中山大學海洋生物研究所博士論文。台灣，中華民國。
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