發(fā)布時(shí)間：2018-05-13 10:28

  本文選題：普通小麥 + 鹽脅迫�。� 參考：《沈陽(yáng)農(nóng)業(yè)大學(xué)》2016年博士論文

【摘要】：小麥?zhǔn)俏覈?guó)第二大口糧作物,其產(chǎn)量和品質(zhì)與國(guó)家糧食安全和人民生活水平息息相關(guān)。鹽脅迫是小麥生產(chǎn)的主要限制因素之一,在轉(zhuǎn)錄和翻譯水平上嚴(yán)重影響了小麥的光合作用、呼吸代謝、水分狀況和各種酶的變化等生理生化過(guò)程。因此在分子水平上闡明小麥對(duì)鹽脅迫的響應(yīng)機(jī)理,鑒定耐鹽關(guān)鍵基因,分析相關(guān)蛋白的修飾水平,對(duì)小麥抗逆育種具有重要的意義。試驗(yàn)以耐鹽小麥品種青麥6號(hào)和鹽敏感材料中國(guó)春為材料,對(duì)人工生長(zhǎng)箱培養(yǎng)1周的小麥幼苗進(jìn)行鹽脅迫處理,利用Illumina測(cè)序平臺(tái)對(duì)鹽脅迫前后6、12、24和48h的小麥根系進(jìn)行高通量測(cè)序,利用生物信息學(xué)方法對(duì)測(cè)序結(jié)果進(jìn)行系統(tǒng)對(duì)比研究,并對(duì)部分同源基因響應(yīng)鹽脅迫時(shí)的差異表達(dá)進(jìn)行了分析。同時(shí),我們利用LC-MS/MS聯(lián)用分析技術(shù)和生物信息學(xué)方法分析小麥乙酰化和琥珀�；�蛋白質(zhì)參與調(diào)控的生理進(jìn)程進(jìn)行了研究。主要結(jié)果如下：1.從全基因組水平上研究了小麥根系響應(yīng)鹽脅迫的基因表達(dá)模式,鑒定到36804個(gè)差異表達(dá)基因,青麥6號(hào)和中國(guó)春鹽脅迫前后基因表達(dá)存在顯著差異。處理前青麥6號(hào)和中國(guó)春大約55.4%和50.1%的部分同源基因之間差異表達(dá),而鹽脅迫后其比例上升到62.6%和59.9%,其差異表達(dá)模式通過(guò)實(shí)時(shí)定量PCR得到了驗(yàn)證。通過(guò)分析鹽脅迫后的上調(diào)表達(dá)的基因發(fā)現(xiàn)6h和12h鹽脅迫處理后青麥6號(hào)上調(diào)基因數(shù)目少于中國(guó)春,但48h后顯著高于中國(guó)春,下調(diào)表達(dá)基因的表達(dá)規(guī)律正好相反。基因GO富集分析顯示,青麥6號(hào)在"cell growth"、"response to ABA stimulus"、"potassium ion transport"、"establishment of localization"和"response to salt stress"等代謝途徑顯著富集,而中國(guó)春在“cell death”、"jasmonic acid biosynthetic process"等代謝途徑富集顯著。2.利用結(jié)構(gòu)域搜索的方法,在小麥基因組中注釋到3718個(gè)轉(zhuǎn)錄因子基因,分布在51個(gè)基因家族,其中1583個(gè)在鹽脅迫下表現(xiàn)出顯著差異表達(dá)。脅迫響應(yīng)轉(zhuǎn)錄因子在不同家族中分布不均衡,差異表達(dá)轉(zhuǎn)錄因子最多的7個(gè)家族(Myb、bZIP、bHLH、NAC、 C2H2、AP2/ERF和WRKY)占所有差異表達(dá)轉(zhuǎn)錄因子總數(shù)的47%。表達(dá)模式聚類(lèi)分析顯示,差異表達(dá)的轉(zhuǎn)錄因子可以聚類(lèi)到兩個(gè)群,分為20種不同的表達(dá)模式。其中第8群和第9群(160個(gè)轉(zhuǎn)錄因子)中,青麥6號(hào)鹽響應(yīng)轉(zhuǎn)錄因子表達(dá)豐度高于中國(guó)春。3.利用生物信息學(xué)方法,將小麥根系鹽脅迫響應(yīng)基因定位到IWGSC發(fā)布小麥參考基因組上,發(fā)現(xiàn)其在染色體上的分布并不是隨機(jī)排列的,而是很多鹽脅迫相關(guān)基因成簇分布。通過(guò)預(yù)測(cè),我們一共鑒定到142個(gè)鹽脅迫基因基因簇,并且發(fā)現(xiàn)它們?cè)贏、B、D部分同源群上部分不均勻,其中D基因組(57)中比A(36)和B(49)具有更高的鹽響應(yīng)熱點(diǎn)。另外通過(guò)基因注釋發(fā)現(xiàn),同一基因簇中的串聯(lián)基因編碼相類(lèi)似的蛋白,但是其響應(yīng)模式具有明顯差異,表明復(fù)制是導(dǎo)致鹽響應(yīng)的熱點(diǎn)區(qū)形成的原因之一,但其功能可能發(fā)生了分化。4.對(duì)小麥賴氨酸乙酰化和琥珀�；�之間的重疊進(jìn)行第一個(gè)全面分析,發(fā)現(xiàn)在277個(gè)蛋白質(zhì)中有416個(gè)特異的乙�；�修飾位點(diǎn),在173個(gè)蛋白中有330個(gè)特異的琥珀�；�修飾位點(diǎn)。根據(jù)GO注釋、KEGG途徑和結(jié)構(gòu)域富集結(jié)果進(jìn)行綜合分析,表明小麥的乙酰化和琥珀�；�修飾作用可以對(duì)細(xì)胞內(nèi)的各種代謝途徑與過(guò)程進(jìn)行調(diào)節(jié)與控制。試驗(yàn)結(jié)果還表明有26個(gè)參與光合作用和卡爾文循環(huán)的蛋白質(zhì)發(fā)生了這2種賴氨酸�；�修飾,21個(gè)琥珀酰賴氨酸位點(diǎn)有相同的乙�；�位點(diǎn),33個(gè)蛋白的乙酰化和琥珀�；�發(fā)生變性,占琥珀酰化的蛋白質(zhì)的7.9%,表明小麥琥珀�；�和乙酰化之間存在重疊。這33種蛋白中的7種酶參與了卡爾文循環(huán),這表明這兩種類(lèi)型的修飾可能在調(diào)節(jié)光合作用和碳固定代謝過(guò)程中發(fā)揮重要作用。本研究為探索在小麥及所有植物的賴氨酸乙�；�和琥珀酰化的生理作用提供了重要參考。
[Abstract]:Wheat is the second largest grain crop in China. Its yield and quality are closely related to national food security and people's living standard. Salt stress is one of the main limiting factors for wheat production. The physiological and biochemical processes, such as photosynthesis, respiration, water status and changes of various enzymes, are seriously affected at the level of transcription and translation. It is important to clarify the response mechanism of wheat to salt stress at the molecular level, identify the key salt tolerant genes and analyze the modification level of the related proteins, which is of great significance to the resistance breeding of wheat. The salt stress of the wheat seedlings with salt tolerant wheat variety green wheat 6 and salt sensitive material for 1 weeks in the artificial growth box was carried out in the experiment. The Illumina sequencing platform was used to sequence the wheat root system of 6,12,24 and 48h before and after salt stress, and the sequencing results were systematically compared with the bioinformatics method, and the differential expression of some homologous genes in response to salt stress was analyzed. At the same time, we used LC-MS/MS combined analysis technology and biological information. The physiological processes of wheat acetylation and succinylated protein were studied. The main results were as follows: 1. the gene expression patterns of wheat root response to salt stress were studied at the whole genome level, and 36804 differentially expressed genes were identified, the expression of gene expression in green wheat 6 and spring salt stress in China was significant. Differences. There was a difference in the expression of some homologous genes between 55.4% and 50.1% of green wheat 6 and China spring before treatment, while the proportion of salt stress increased to 62.6% and 59.9%, and its differential expression pattern was verified by real-time quantitative PCR. By analyzing the up-regulated gene of salt stress, it was found that the number of green wheat 6 was up after 6h and 12h stress treatment. The number of genes was less than that of spring in China, but after 48h, it was significantly higher than that of China spring. The expression of down regulated genes was the opposite. The gene GO enrichment analysis showed that the metabolic pathways such as "cell growth", "response to ABA stimulus", "potassium ion transport", "establishment ion" and "establishment" and other metabolic pathways were significantly enriched. In the spring of China, in the "cell death", "jasmonic acid biosynthetic process" and other metabolic pathways, a significant.2. use domain search method was used. In the wheat genome, 3718 transcription factor genes were annotated and distributed in 51 gene families. 1583 of them showed significant differential expression under salt stress. The stress response transcription factors were not The 7 families (Myb, bZIP, bHLH, NAC, C2H2, AP2/ERF and WRKY) with the most differentially expressed transcriptional factors (Myb, bZIP, C2H2, AP2/ERF and WRKY) accounted for the 47%. expression pattern of all the total differential expression transcription factors. The differentially expressed transcription factors could be clustered into two groups and divided into 20 different expression patterns. Among them, eighth and ninth groups (16 In the 0 transcription factors, the expression abundance of the salt response transcription factor of the green wheat 6 was higher than that of the Chinese Spring.3. bioinformatics method. The salt stress response gene of wheat root system was located to IWGSC to publish the wheat reference genome, and it was found that the distribution of the genes on the chromosomes was not arranged randomly, but a lot of salt stress related genes were distributed. Over prediction, we have identified 142 salt stress gene clusters and found that they are partially inhomogeneous on A, B, and D homologous groups, and the D genome (57) has a higher salt response than A (36) and B (49). There is a distinct difference, indicating that replication is one of the reasons for the formation of salt response hot spots, but its function may have occurred in the first comprehensive analysis of the overlap between the acetylation and succinylation of wheat lysine, and found that there were 416 specific acetylation modification sites in the 277 proteins and 330 of the 173 proteins in the 277 proteins. A specific succinylation site. Based on the GO annotation, the KEGG pathway and the domain accumulation results, the results showed that the acetylation and succinylation of wheat could regulate and control the various metabolic pathways and processes in the cells. The results also showed that 26 eggs involved in photosynthesis and the Calvin cycle. These 2 lysine acylation modifications occur in white matter, 21 succinyl lysine sites have the same acetylation sites, 33 proteins have acetylation and succinylation, accounting for 7.9% of succinylated proteins, indicating that there is a overlap between succinylation and acetylation of the wheat. The 7 enzymes of the 33 proteins participate in the Calvin cycle. These two types of modification may play an important role in regulating photosynthesis and carbon immobilization. This study provides an important reference for the exploration of the physiological role of lysine acetylation and succinylation in wheat and all plants.

【學(xué)位授予單位】：沈陽(yáng)農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：S512.1
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本文編號(hào)：1882772