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研究生:周逸潔
研究生(外文):Yi-Jia Chou
論文名稱:水稻醛酮還原酶 AKR5環境逆境耐受性機制之研究
論文名稱(外文):水稻醛酮還原酶 AKR5環境逆境耐受性機制之研究
指導教授:黃文理黃文理引用關係
指導教授(外文):Wen-Lii Huang
學位類別:碩士
校院名稱:國立嘉義大學
系所名稱:農藝學系研究所
學門:農業科學學門
學類:一般農業學類
論文種類:學術論文
畢業學年度:104
語文別:中文
中文關鍵詞:水稻醛酮還原酶
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  • 被引用被引用:3
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本論文為延續本研究室針對水稻醛酮糖還原酶 (aldo-keto reductase, AKR) 在基因剔除突變系 (KoAKR5)、過量表現轉殖系 (OxAKR5)進行相關生理分析、基因表現,以及利用次世代定序 (next-generation sequencing, NGS) 方法進行水稻基因體組分析,與其基因功能性之探討。
本論文試驗比較過量表現OsAKR5、KoAKR5 (M16610) 及WT (TNG67) 於4℃、0.2M sorbitol 逆境下之光合效率,結果顯示光合作用效率為過量表現轉殖系優於WT及KoAKR5。利用3,3’-diaminobenzidine (DAB) 及nitroblue tetrazolium (NBT) 進行染色,結果顯示過量表現系於逆境下的呈色面積均較WT及KoAKR5少,顯示過量表現OsAKR5具減少逆境引起之活性氧族含量。本論文進一步利用RT-PCR分析WT及過量表現OsAKR5於逆境下的基因表現,結果顯示OsAKR5基因在滲透壓逆境下,其表現量增加,而KoAKR5則偵測不到其表現量之表現。另外,在NGS的結果中,將生長至3-4葉齡的水稻幼苗處理0.2M sorbitol 3天,建立cDNA資料庫,經基因鑑定後,測序資料庫共有82633筆基因註釋。比較其表現差異(fold change >2)0.2M的sorbitol滲透壓逆境下,OxAKR5和WT共有414筆資料,並透過RT-PCR證實部分受逆境調控的基因表現。這些結果表明,OsAKR5在水稻發育過程中,於對非生物逆境之耐受性中為一關鍵角色。
This study continued our previous studies on rice aldo-keto reductase on the OsAKR5 knock out mutant (KoAKR5), overexpressed transgenic lines (OxAKR5) for further functional and physiological analysis. Also, we used NGS technology to investigate genome-wide gene expression in response to abiotic stress.
In our study, we observed the phenotypic between these three lines under the cold and osmotic stresses. Also, we compared the chlorophyll fluorescence (Fv/Fm), the result indicated that the overexpressed transgenic lines maintain higher Fv/Fm than wild type and knock out line. On the other hand, we found that OxAKR5s transformants reduced H2O2and O2- content under both temperature stress and osmotic stress by 3,3’-diaminobenzidine (DAB) and nitroblue tetrazolium (NBT) staining. These result indicated that OsAKR5 could play an important role on reducing H2O2and O2- under the abiotic stress. Also, we analyzed the gene expression of KoAKR5, OxAKR5 and WT under the osmotic stress by RT-PCR. The result indicated that the gene expression of OxAKR5 would induce under the osmotic stress, but cannot detect in KoAKR5 mutant.
As for the result of NGS, we constructed cDNA libraries for the whole plant of rice plants grown under 0.2M sorbitol condition for 3 days. Sequencing the libraries resulted in identification of 82,633 annotated genes. A comparison of abundances revealed 414 transcripts that were differentially expressed (fold change >2) due to the OxAKR5 and WT under the 0.2 M sorbitol condition. Responses by different abiotic stress genes were confirmed by RT-PCR. These results indicate that AKR5 plays a pivotal role in stress tolerance and plant development in rice.
中文摘要................................................................................... I
英文摘要................................................................................... II
目錄.......................................................................................... IV
表目錄....................................................................................... V
圖目錄......................................................................................VII
一、前言................................................................................... 1
二、前人研究.............................................................................. 4
三、材料方法............................................................................ 28
四、結果.................................................................................... 49
五、討論.................................................................................... 58
六、參考文獻............................................................................ 67
七、附錄.................................................................................. 113
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