本文選題：敖漢苜蓿 + 小花　； 參考：《中國農(nóng)業(yè)大學(xué)》2017年博士論文

【摘要】：硼是紫花苜蓿必需的微量元素之一,它能促進(jìn)紫花苜蓿生殖器官的正常發(fā)育,有利于開花結(jié)實,對提高苜蓿種子產(chǎn)量和質(zhì)量的作用不可估量。苜蓿對硼的需求在適量和過量之間的界限很小,施用不當(dāng)會使苜蓿中毒從而降低種子產(chǎn)量。因此開展紫花苜蓿硼營養(yǎng)機理以及紫花苜蓿生殖器官響應(yīng)硼脅迫的機制研究,有助于實際生產(chǎn)過程中合理使用硼肥,有效提高苜蓿種子產(chǎn)量。本論文在實際生產(chǎn)實驗的基礎(chǔ)上,以敖漢苜蓿小花和種子為實驗材料,應(yīng)用蛋白質(zhì)組學(xué)和代謝組學(xué)技術(shù),研究硼脅迫下蛋白質(zhì)及小分子代謝產(chǎn)物的變化及代謝途徑,探索紫花苜蓿生殖器官響應(yīng)硼脅迫的應(yīng)答機制。主要研究內(nèi)容如下:1.選用0、0.04、0.08、0.12和0.16%濃度的硼,對生長第二年的敖漢苜蓿進(jìn)行施肥實驗。結(jié)果表明,施硼肥有效提高紫花苜蓿小花花粉數(shù)量、花粉活力、種子產(chǎn)量、產(chǎn)量構(gòu)成因子及種子質(zhì)量�？傮w上,0.08%濃度處理組的效果較好,增產(chǎn)效果顯著。而0.12%處理組存在落花現(xiàn)象,從而降低種子產(chǎn)量。當(dāng)噴施濃度達(dá)0.16%時,已產(chǎn)生毒害作用。因此選擇0.00%濃度作為缺硼處理,0.08%濃度作為最適施肥量,0.16%濃度作為高硼處理進(jìn)行下一步研究。2.測定了 0%(缺硼)、0.08%(對照)和0.16%(高硼)硼處理的敖漢苜蓿小花與種子的抗氧化酶活性、MDA、脯氨酸和可溶性糖含量。測定結(jié)果表明,在缺硼脅迫下敖漢苜蓿小花與種子的抗氧化酶活性、MDA和脯氨酸含量均有不同程度的下降。在高硼處理下敖漢苜蓿種子抗氧化酶活性和MDA含量增加,而脯氨酸和可溶性糖含量下降。高硼脅迫對敖漢苜蓿小花的影響不大,但對種子造成一定的影響。3.運用2-DE技術(shù)對0%、0.08%和0.16%硼處理的敖漢苜蓿小花與種子蛋白質(zhì)進(jìn)行分離,并用PDQuest圖像軟件、LC-MS/MS質(zhì)譜技術(shù)、Mascot軟件和Uniprot數(shù)據(jù)庫進(jìn)行分析鑒定。在缺硼脅迫下從敖漢苜蓿小花中成功鑒定出12個差異蛋白,主要參與碳水化合物代謝和脂質(zhì)代謝。從種子中成功鑒定出11個差異蛋白,主要參與碳水化合物代謝和氨基酸代謝。在高硼脅迫下從敖漢苜蓿小花中共篩選出14個差異蛋白,主要參與碳水化合物代謝、氨基酸代謝、脂質(zhì)代謝和次生代謝。從種子中共篩選出11個差異蛋白,主要參與了氨基酸代謝、碳水化合物代謝和脂質(zhì)代謝。敖漢苜蓿小花酰基蛋白硫酯酶在缺硼和高硼脅迫下均下調(diào)表達(dá),敖漢苜蓿種子NAD依賴的醛脫氫酶家族蛋白在缺硼和高硼脅迫下均上調(diào)表達(dá)。NAD(P)結(jié)合羅斯曼折疊蛋白在高硼處理的紫花苜蓿小花與種子中均表達(dá),是紫花苜蓿響應(yīng)硼毒害的關(guān)鍵調(diào)控蛋白。4.利用GC-MS技術(shù),對缺硼和高硼脅迫下敖漢苜蓿小花與種子進(jìn)行代謝物譜分析。采用OPLS-DA模型第一主成分的VIP值(閾值1)結(jié)合單維統(tǒng)計(t檢驗)的p值(閾值p0.05)尋找差異性表達(dá)代謝物。在缺硼脅迫處理組中共篩選到50個差異物質(zhì),主要涉及碳水化合物代謝和氨基酸代謝;從種子中共篩選到13個差異物質(zhì),主要參與了氨基酸代謝。在高硼脅迫下從敖漢苜蓿小花中共篩選到97個差異物質(zhì),主要參與氨基酸代謝、碳水化合物代謝、脂質(zhì)代謝、嘌呤代謝和嘧啶代謝;從種子中共篩選并定性到16個差異物質(zhì),主要參與氨基酸代謝。5.綜合以上分析發(fā)現(xiàn),在硼脅迫下紫花苜蓿小花與種子主要通過調(diào)控氨基酸代謝、碳水化合物代謝、脂質(zhì)代謝和三羧酸循環(huán)等代謝途徑來適應(yīng)硼脅迫,從而適應(yīng)硼營養(yǎng)缺乏或過量環(huán)境。
[Abstract]:Boron is one of the essential trace elements of alfalfa. It can promote the normal development of the reproductive organs of alfalfa, be beneficial to the flowering and fruiting, and immeasurable effect on improving the yield and quality of alfalfa seeds. The limit of the amount and excess of alfalfa to boron is very small. The mechanism of boron nutrition in alfalfa and the mechanism of boron stress in the reproductive organs of alfalfa are studied. It is helpful to use boron fertilizer in the actual production process and improve the yield of Alfalfa effectively. On the basis of actual production experiments, the experimental materials of the Aohan alfalfa flower and seed seed are used as the experimental materials, and the proteomics and metabolic groups are applied. To study the changes and metabolic pathways of protein and small molecule metabolites under boron stress and explore the response mechanism of the reproductive organs of Alfalfa in response to boron stress. The main contents are as follows: 1. the application of 0,0.04,0.08,0.12 and 0.16% concentration of boron to the growth of Alfalfa for second years was carried out. The results showed that the application of boron fertilizer was effective. Increase the number of pollen, pollen vigor, seed yield, yield components and seed quality of alfalfa. In general, the effect of 0.08% concentration treatment group is better, and the effect of increasing yield is remarkable. The 0.12% treatment group has the phenomenon of falling flower, which reduces the seed yield. When the spraying concentration reaches 0.16%, it has the toxic effect. So 0% concentration is chosen. For boron deficiency treatment, 0.08% concentration as the optimum fertilizer amount, 0.16% concentration as high boron treatment, the antioxidant enzyme activity, MDA, proline and soluble sugar content of Alfalfa small flowers and seeds treated by.2., 0% (boron deficiency), 0.08% (control) and 0.16% (Gao Peng) boron treated alfalfa were measured. The results showed that the alfalfa under boron deficiency stress The activity of antioxidant enzymes and the content of MDA and proline decreased in different degrees. Under the high boron treatment, the antioxidant enzyme activity and MDA content of alfalfa seeds increased, while the content of proline and soluble sugar decreased. The effect of high boron stress on the alfalfa small flowers was not significant, but the effect of.3. on the seeds of the seeds was a certain effect on the use of 2-DE technology. 0%, 0.08% and 0.16% boron treated alfalfa flowers and seed proteins were separated and analyzed by PDQuest image software, LC-MS/MS mass spectrometry, Mascot software and Uniprot database. Under boron deficiency stress, 12 differentially identified proteins were successfully identified from the alfalfa small flowers, mainly involved in carbohydrate metabolism and lipid. Metabolism. 11 differentially identified proteins were identified from the seeds, mainly involved in carbohydrate metabolism and amino acid metabolism. Under high boron stress, 14 differential proteins were screened from the alfalfa small flower, mainly involved in carbohydrate metabolism, amino acid metabolism, lipid metabolism and secondary metabolism. 11 differential proteins were screened from the seeds. The amino acid metabolism, carbohydrate metabolism and lipid metabolism were involved. Ohan alfalfa protein thioesterase was downregulated under boron deficiency and high boron stress. The NAD dependent aldehyde dehydrogenase family protein up regulated the expression of.NAD (P) combined with Rosman foldable protein in high boron treatment under boron deficiency and high boron stress. Alfalfa was expressed in both small flowers and seeds. It was the key regulatory protein of alfalfa to respond to boron toxicity,.4. using GC-MS technology to analyze the metabolite spectrum of the small flowers and seeds of Alfalfa under boron deficiency and high boron stress. The VIP value of the first principal component of the OPLS-DA model (threshold 1) was combined with the p value of the single dimension statistical (t test) to find the difference. A total of 50 different substances, including carbohydrate metabolism and amino acid metabolism, were mainly involved in the metabolism of carbohydrates and amino acids. 13 different substances were selected from the seeds and mainly involved in the metabolism of amino acids. Under the stress of high boron, 97 different substances were selected from the alfalfa small flower, and they were mainly involved in the amino acid generation. Metabolism of carbohydrates, lipid metabolism, purine metabolism, and pyrimidine metabolism; screening and qualitative analysis from the seeds to 16 different substances, mainly involved in the amino acid metabolism.5. synthesis analysis found that under boron stress, alfalfa small flowers and seeds are mainly regulated by amino acid metabolism, carbohydrate metabolism, lipid metabolism, and three carboxylic acids. Ring and other metabolic pathways are adapted to boron stress, so as to adapt to boron deficiency or excessive environment.
【學(xué)位授予單位】：中國農(nóng)業(yè)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：S541.9

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