本文選題：植物代謝組學 + 植物激素��； 參考：《中國農(nóng)業(yè)大學》2017年博士論文

【摘要】：植物代謝物種類繁多、結構迥異,是植物生命活動重要的體現(xiàn)者。對植物代謝物進行系統(tǒng)深入的研究,挖掘代謝物背后蘊含的生物學意義,有利于加強對植物生命活動現(xiàn)象、本質(zhì)與規(guī)律的理解,對增強植物對不良環(huán)境的抗性、提高作物產(chǎn)量與品質(zhì)具有重要意義。本論文應用基于高分辨質(zhì)譜的植物代謝組學技術,建立了 6種植物激素與40種游離氨基酸的檢測方法;結合基因表達分析,研究了茉莉酸對擬南芥代謝組的影響;對大芻草-玉米代謝組進行了比較研究,并對玉米馴化過程中的代謝物變異進行了初步的遺傳解析。植物激素檢測面臨著含量低、性質(zhì)不穩(wěn)定、基質(zhì)復雜的困難,本研究應用液-液萃取與同位素稀釋技術,建立了 6種植物激素的檢測方法。結果表明,液-液萃取提供了較高的提取效率,同位素稀釋校正了樣品分析過程中的偏差,超高效液相色譜與高分辨質(zhì)譜保證了方法的特異性與靈敏度。對方法的準確度、精度、靈敏度、線性和回收率進行評價,表明該方法滿足實際分析的需要,同時使用該方法對擬南芥中植物激素的時間與空間分布進行了初步的分析。應用柱前衍生、反相超高效液相色譜與高分辨質(zhì)譜,建立了 40種游離氨基酸的檢測方法。在解決了氨基酸極性較強、缺少顯色基團問題的同時,提高了分離效率與檢測靈敏度。對方法的特異性、線性、靈敏度、回收率和重現(xiàn)性進行驗證,該方法具有快速、高效和高靈敏度的優(yōu)點。結合主成分分析,對多種蔬菜中的游離氨基酸含量進行了比較。結果表明,不同蔬菜中游離氨基酸的種類大致相同,但其含量存在顯著差異;該方法適用于基質(zhì)復雜的植物材料,也可以擴展到果汁、水果等其它相關樣品的分析。對擬南芥野生型、茉莉酸合成缺失突變體opr3以及茉莉酸處理后的opr3突變體進行了代謝組分析,一共檢測到超過10000個化合物離子,其中2000多個離子信號在不同的基因型或處理組間存在顯著差異。統(tǒng)計分析結果表明擬南芥野生型與突變體opr3存在明顯的代謝物差異,外源茉莉酸處理能使突變體中部分代謝物逐漸恢復到野生型的水平。本實驗還建立了一個差異代謝物數(shù)據(jù)庫,包含了 174個代謝物的保留時間、母離子與特征二級碎片的精確質(zhì)量,以及109個代謝物的化學結構。對鑒定到的差異代謝物進行了聚類分析與通路分析,結果表明硫代葡萄糖苷與色氨酸代謝、脂類代謝、氨基酸、小肽等都受到了茉莉酸的調(diào)節(jié)。使用熒光定量反轉錄PCR對相關通路中的重要基因進行了表達量的檢測,驗證了代謝組的結果,這些結果揭示了植物激素與植物代謝物之間的相互作用與調(diào)節(jié)網(wǎng)絡,加強了我們對茉莉酸生物功能的理解。比較了 30個大芻草和40個玉米品系的代謝組,檢測了 624份玉米與大芻草重組自交系群體中的代謝物,并對挖掘到的重要代謝物進行靶標分析。結果表明玉米與大芻草代謝組存在差異,鑒定到97個玉米與大芻草間分化的代謝物。在重組自交系群體中,鑒定到了 299個有QTL定位結果的代謝物,這些代謝物種類各異,參與了多條重要的代謝通路。后續(xù)對重要差異代謝物的遺傳解析,并克隆相關基因?qū)τ衩椎倪z傳改良具有重要的指導意義。
[Abstract]:Plant metabolites are diverse and different in structure. It is an important embodiment of plant life activities. The systematic and in-depth study of plant metabolites and the biological significance behind the metabolites are beneficial to the understanding of plant life activities, the understanding of the nature and the law, the resistance to the adverse environment and the increase of crop yield. In this paper, 6 kinds of plant hormones and 40 kinds of free amino acids were established based on high resolution mass spectrometry, and the effects of jasmonic acid on the metabolic group of Arabidopsis were studied in combination with gene expression analysis. Preliminary genetic analysis of metabolite variation in the process is carried out. Plant hormone detection is faced with low content, unstable properties and complex matrix. In this study, liquid liquid extraction and isotope dilution technique were used to establish 6 methods for detecting plant hormones. The results showed that liquid liquid extraction provided high extraction efficiency and isotopic dilution. The accuracy, accuracy, sensitivity, linearity and recovery of the method were evaluated by ultra high performance liquid chromatography and high resolution mass spectrometry. The method met the needs of actual analysis, and the time and space of plant hormones in Arabidopsis were used. 40 kinds of free amino acid detection methods were established by using pre column derivatization, reversed phase ultra high performance liquid chromatography and high resolution mass spectrometry. The separation efficiency and detection sensitivity were improved. The specificity, linearity, sensitivity and recovery of the method were improved. This method has the advantages of rapid, high efficiency and high sensitivity. The content of free amino acids in various vegetables is compared with principal component analysis. The results show that the species of free amino acids in different vegetables are roughly the same, but the content of free amino acids is significantly different. This method is suitable for complex matrix plants. The material can also be extended to the analysis of other related samples, such as fruit juice, fruit, and other related samples. The metabolic group analysis was carried out on the Arabidopsis wild type, the jasmonic acid synthesis deletion mutant opr3 and the opr3 mutant after jasmonic acid treatment. More than 10000 compound ions were detected, of which more than 2000 ion signals were stored in different genotypes or treatment groups. The statistical analysis showed that there was a significant difference in metabolites between the Arabidopsis wild type and the mutant opr3. Exogenous jasmonic acid treatment could make some metabolites in the mutant gradually recover to the level of the wild type. A different metabolite database was established in this experiment, including the retention time of 174 metabolites, the mother ion and the mother ion. The exact mass of the two grade fragments and the chemical structure of the 109 metabolites were characterized by cluster analysis and pathway analysis of the identified metabolites. The results showed that glucosinolates and tryptophan metabolism, lipid metabolism, amino acids, small peptides were regulated by jasmonic acid. The fluorescence quantitative reverse transcriptional PCR was used in the related pathways. The results of the metabolic group were verified by the detection of the important genes, which revealed the interaction and regulation network between plant hormones and plant metabolites, and enhanced our understanding of the biological function of jasmonic acid. The metabolic groups of 30 rice and 40 maize lines were compared, and 624 corn and rice were detected. The metabolites in the population of the inbred lines were reorganized and the important metabolites were analyzed. The results showed that there were differences in the metabolic group between the maize and the rice. The metabolites differentiated between 97 maize and the rice were identified. In the recombinant inbred population, 299 metabolites with QTL location results were identified, and the metabolites of these metabolites were different. The genetic analysis of important differential metabolites and cloning of related genes have important guiding significance for the genetic improvement of maize.
【學位授予單位】：中國農(nóng)業(yè)大學
【學位級別】：博士
【學位授予年份】：2017
【分類號】：Q945

【參考文獻】

相關期刊論文 前1條

1 郭繼芬;張紹東;孟繁華;趙毅民;;液相色譜-串聯(lián)質(zhì)譜法測定大鼠腦透析液中CTN986及其脫糖產(chǎn)物的含量[J];高等學�；瘜W學報;2009年08期

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