本文選題：小麥　切入點(diǎn)：NAC　出處：《華中科技大學(xué)》2016年碩士論文

【摘要】：干旱、高鹽以及極端溫度等不利因素嚴(yán)重影響植物的正常生長(zhǎng)發(fā)育,使農(nóng)作物的產(chǎn)量下降、品質(zhì)降低。植物在長(zhǎng)期的進(jìn)化過程中,形成了一系列調(diào)控機(jī)制以適應(yīng)或抵御不利環(huán)境的影響,轉(zhuǎn)錄因子調(diào)控基因表達(dá)就是其中之一。轉(zhuǎn)錄因子可以通過與相應(yīng)的順式作用元件結(jié)合,調(diào)控相關(guān)基因的時(shí)空表達(dá),響應(yīng)不同逆境脅迫。近年來植物特有且為最大的轉(zhuǎn)錄因子家族之一的NAC(NAM,ATAF與CUC)在抗逆功能方面的研究報(bào)道越來越多,但大部分研究工作都集中在模式植物擬南芥和水稻上,而關(guān)于小麥NAC轉(zhuǎn)錄因子的功能研究還鮮有報(bào)道。本研究在實(shí)驗(yàn)室前期工作基礎(chǔ)上,分析了小麥NAC轉(zhuǎn)錄因子家族成員Ta NAC8B基因的表達(dá)模式和轉(zhuǎn)錄激活活性;通過根癌農(nóng)桿菌介導(dǎo)的遺傳轉(zhuǎn)化法獲得了Ta NAC8B過表達(dá)的轉(zhuǎn)基因煙草株系,并研究了其抗逆功能。主要研究成果如下:(1)通過模擬不同逆境脅迫,用實(shí)時(shí)熒光定量PCR方法檢測(cè)了Ta NAC8B基因的表達(dá)變化,發(fā)現(xiàn)PEG6000處理可顯著上調(diào)Ta NAC8B的表達(dá),而Na Cl、ABA和低溫等脅迫處理下Ta NAC8B表達(dá)量沒有顯著變化。(2)構(gòu)建p GBKT7-Ta NAC8B真核表達(dá)載體,通過酵母自激活實(shí)驗(yàn),發(fā)現(xiàn)Ta NAC8B具有轉(zhuǎn)錄激活活性,且轉(zhuǎn)錄激活活性區(qū)域位于Ta NAC8B C-端。(3)利用根癌農(nóng)桿菌介導(dǎo)的遺傳轉(zhuǎn)化技術(shù),成功將Ta NAC8B轉(zhuǎn)入煙草并獲得陽性植株。通過對(duì)T1代種子的模擬逆境脅迫處理下的根長(zhǎng)實(shí)驗(yàn),發(fā)現(xiàn)Ta NAC8B過表達(dá)植株表現(xiàn)出較強(qiáng)的抗逆性。進(jìn)一步通過對(duì)幼苗的干旱脅迫處理,觀察表型以及測(cè)定相關(guān)生理指標(biāo),發(fā)現(xiàn)Ta NAC8B過表達(dá)植株表現(xiàn)出較強(qiáng)的抗旱性能,這與Ta NAC8B基因表達(dá)模式分析和根長(zhǎng)實(shí)驗(yàn)結(jié)果一致。綜上所述,Ta NAC8B轉(zhuǎn)錄因子可以響應(yīng)干旱逆境脅迫處理,過表達(dá)Ta NAC8B植株表現(xiàn)出較強(qiáng)的干旱脅迫耐受性,這為進(jìn)一步研究Ta NAC8B的基因功能奠定了基礎(chǔ)。
[Abstract]:Drought, high salt, extreme temperature and other adverse factors seriously affect the normal growth and development of plants, resulting in lower crop yields and lower quality. Plants have evolved over a long period of time. A series of regulatory mechanisms have been formed to adapt to or resist adverse environmental effects, and transcription factor regulation gene expression is one of them. Transcription factors can regulate the expression of related genes in time and space by combining with the corresponding cis-acting elements. In recent years, there are more and more studies on stress resistance of NACU NAMMAF and CUC, one of the largest transcription factor families, but most of the research work is focused on Arabidopsis thaliana and rice. However, there are few reports on the function of wheat NAC transcription factors. Based on the previous work in laboratory, the expression pattern and transcriptional activation activity of Ta NAC8B gene, a member of wheat NAC transcription factor family, were analyzed. Transgenic tobacco lines with Ta NAC8B overexpression were obtained by Agrobacterium tumefaciens mediated genetic transformation, and their stress resistance was studied. The expression of Ta NAC8B gene was detected by real-time fluorescence quantitative PCR. It was found that PEG6000 treatment could significantly upregulate the expression of Ta NAC8B, while the expression of Ta NAC8B did not change significantly under NaCl NAC8B and low temperature stress.) the eukaryotic expression vector of p GBKT7-Ta NAC8B was constructed. Through yeast self-activation experiment, we found that Ta NAC8B has transcriptional activation activity, and the transcriptional activation region is located at the C- terminal of Ta NAC8B, using Agrobacterium tumefaciens mediated genetic transformation. Ta NAC8B was successfully transferred into tobacco and positive plants were obtained. Through the experiment of root length of T1 generation seeds under simulated stress stress, it was found that Ta NAC8B overexpression plants showed strong resistance to stress. The phenotypic and physiological indexes were observed, and it was found that the plants with Ta NAC8B overexpression showed strong drought resistance. These results are consistent with the analysis of Ta NAC8B gene expression pattern and the results of root length experiment. In conclusion, Ta NAC8B transcription factors can respond to drought stress stress, and over-expressed Ta NAC8B plants exhibit strong tolerance to drought stress. This lays a foundation for further studying the gene function of Ta NAC8B.
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：Q943.2;S512.1

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