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標題: | 阿拉伯芥中miR160/AUXIN RESPONSE FACTOR (ARF)10、ARF16及ARF17調節熱逆境反應機制 miR160 and AUXIN RESPONSE FACTOR10, 16, and 17 affects heat responses in Arabidopsis |
作者: | Chia-Chia Kuo 郭家嘉 |
指導教授: | 鄭石通(Shih-Tong Jeng) |
關鍵字: | 阿拉伯芥,miR160,熱逆境,Auxin Response Factor (ARFs),Heat Shock Protein (HSPs),耐熱性, Arabidopsis,miR160,Auxin,Auxin Response Factor (ARFs),Heat Shock Protein (HSPs),Thermotolerance, |
出版年 : | 2015 |
學位: | 碩士 |
摘要: | 熱逆境會負向影響植物生長發育,甚至造成細胞凋亡,影響植物的生殖與產量。在本研究中,分析熱逆境下的阿拉伯芥的DNA微陣列 (microarray),發現AUXIN RESPONSE FACTOR 10 (ARF10)會受熱調控,ARF10也是miR160的目標基因。實驗室前人在基因表現檢測實驗中也證實熱逆境會誘導miR160及其前驅物基因的表現,進而使其目標基因ARF10、ARF16及ARF17表現量降低。因此,為了研究miR160在熱逆境下扮演的角色,構築大量表現miR160a (160OE) 及抑制miR160功能 (MIM160) 的轉殖株,並以即時定量PCR檢測轉殖株中ARF10、ARF16及ARF17表現。在160OE轉殖株中,目標基因表現被抑制;反之,在MIM160轉殖株中,其目標基因不受miR160調控,而使得ARF10、ARF16及ARF17的表現量可以累積。在熱逆境下,160OE轉殖株的種子發芽率、存活率、下胚軸及花軸發育的部分,相較於野生型,具有較為耐熱的表型;反之,MIM160轉殖株則具有較不耐熱的表型。因此,由這些實驗結果得知,miR160表現量的提升會幫助植株的耐熱性。此外,我們也從ABRC獲得miR160目標基因的T-DNA突變株 (arf10-1、arf16-3及arf17-2) 。在熱逆境下,arf10-1、arf16-3及arf17-2突變株的發芽率及花軸發育,與160OE轉殖株相似,也具有耐熱的表型。並且發現arf16-3及arf17-2突變株,在熱逆境下具有較野生型長的下胚軸表型。這些實驗的結果指出熱逆境下miR160目標基因會負向調控植物生長發育,參與在熱反應的調控中。研究進一步發現,以即時定量PCR檢測,在熱逆境下160OE轉植株會累積較高的熱休克蛋白,包含HSP70B、HSP21、HSP17.6A及HSP17.6?,而在MIM160轉植株中則會有相對低的累積量。綜合實驗結果得知,在高溫逆境下,阿拉伯芥能藉由誘導miR160的表現量上升,抑制其目標基因ARF10、ARF16及ARF17,並影響熱休克蛋白HSP70B、HSP21、HSP17.6A及HSP17.6?的表現,調控植物在熱逆境時的反應。 High temperature negatively affects plant growth and development, and even causes cell death and reduces crop yield. Several heat-related genes obtained from microarray analyses are targets of miRNAs, including AUXIN RESPONSE FACTOR10 (ARF10), the target of miR160. The results of gene expression assay indicated that miR160 and its precursors were induced by heat and its targets, ARF10, ARF 16, and ARF17, were significantly repressed. Hence, transgenic plants overexpressing pre-miR160a(160OE)and expressing miR160 target mimic inhibitor(MIM160)were generated to investigate the roles of miR160 under heat stress. The expression of ARF10, ARF16, and ARF17 was decreased in 160OE plants and increased in MIM160 plants in heat stress. When 160OE plants were treated with heat stress, seed germinations, survival rates, hypocotyl lengths, and rachis growth of 160OE were elevated compared to those of WT. However, these phenotypes in MIM160 plants were worse than WT after heat treatment. These results indicated that increasing miR160 expression could enhance thermotolerant abilities of plants. In addition, arf10, arf16, and arf17 mutants ordered from ABRC also presented better adaption to heat than WT. After heat treatment, the seed germination and inflorescence stem length of arf10, arf16, and arf17 mutants were better and longer, respectively, than those of WT. The arf16 and arf17 mutants also showed the increased hypocotyl lengths compared to WT under heat treatment. These results demonstrated that miR160 targets may negatively regulate the plant development under heat stress. Furthermore, the expression of HSP101, HSP70B, HSP21, HSP17.6A, and HSP17.6II was affected in 160OE and MIM160 plants under heat stress. In conclusion, miR160 and its targets alter the plant development and the HSPs gene expression to regulate plant responses in heat stress. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/54021 |
全文授權: | 有償授權 |
顯示於系所單位: | 植物科學研究所 |
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