本文關(guān)鍵詞： 玉米 油菜素內(nèi)酯 ABA 丙環(huán)唑 ZmBRI1　出處：《中國農(nóng)業(yè)大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：油菜素內(nèi)酯(BRs)是一種重要的甾族類激素,在植物生長發(fā)育過程中起著重要的調(diào)控作用,同時也參與植物對逆境響應(yīng)調(diào)控。論文針對玉米生產(chǎn)中增密或肥料管理不當(dāng)?shù)葘?dǎo)致群體質(zhì)量下降以及非生物逆境如干旱等影響植株生長等生產(chǎn)問題,利用BRs調(diào)節(jié)株型與響應(yīng)逆境調(diào)控效應(yīng),開展BR抑制劑丙環(huán)唑(Pcz)調(diào)節(jié)玉米生長效應(yīng)及其調(diào)控機制,為Pcz在玉米株型塑造上應(yīng)用提供依據(jù);同時,克隆玉米的BR受體基因ZmBRI1,分析其功能,研究了 BR信號表達(dá)響應(yīng)ABA的蛋白質(zhì)調(diào)控機制,為揭示BR與ABA互作響應(yīng)逆境提供理論依據(jù)。主要研究結(jié)果如下:1)Pcz處理顯著抑制玉米中胚軸與胚芽鞘的生長,降低了株高,縮短了葉片和葉鞘長度,減小了葉夾角,同時顯著抑制了葉片和葉鞘細(xì)胞的縱向伸長,促使葉枕細(xì)胞排列由疏松變?yōu)榫o密;降低了玉米中赤霉素(GA)的含量,下調(diào)了GA3ox 表達(dá),上調(diào)了 GA失活的酶GA2ox5和GA2ox8表達(dá),而GA合成酶GA20ox1的表達(dá)呈現(xiàn)先上調(diào)后下調(diào)模式;降低了油菜素內(nèi)酯(BR)含量,但BR合成基因CPD和DWF4的表達(dá)上調(diào),可能是由于反饋調(diào)節(jié);下調(diào)了擴張蛋白EXZR4、EXPA5和木葡聚糖內(nèi)糖基轉(zhuǎn)移酶/水解酶基因XTH1、XET1的表達(dá)。2)利用同源克隆的方法,從玉米B73自交系中獲得了油菜素內(nèi)酯受體蛋白編碼基因ZmBRI1,全長為3369bp,編碼1122個氨基酸;亞細(xì)胞定位分析表明ZmBRI1蛋白定位于細(xì)胞膜上,且在玉米各個組織器官中都有表達(dá);利用轉(zhuǎn)基因技術(shù)將ZmBRI1導(dǎo)入擬南芥BR不敏感突變體bri1-5中,修復(fù)了突變體bri1-5的表型,如植株高度、葉片形態(tài)和果莢大小;修復(fù)了突變體bri1-5的BR信號,如根長、下胚軸和BR合成酶基因CPD、DWF4的表達(dá);在ABA處理條件下,ZmBRI1超表達(dá)株系line2和line5顯著提高了種子的萌發(fā)率和根長,且ABA響應(yīng)基因RS29A、RD29B、ABI5和RAB18表達(dá)量降低,對ABA敏感性減弱。3)利用iTRAQ技術(shù)對野生型Ws、突變體bri1-5和ZmBRI1超表達(dá)株系line5在幼苗生長過程中響應(yīng)ABA的蛋白質(zhì)組進行了分析,共有5479個蛋白被鑒定,其中差異蛋白263個;鑒定出的差異蛋白涉及了初生代謝和次生代謝、能量和運輸、細(xì)胞生長和分裂、蛋白合成和降解、轉(zhuǎn)錄和信號傳導(dǎo)、脅迫響應(yīng)和細(xì)胞結(jié)構(gòu)等多種生物學(xué)途徑;突變體bri1-5對ABA最為敏感,物質(zhì)代謝、能量轉(zhuǎn)化和蛋白合成平衡被打破,脅迫響應(yīng)基因的表達(dá)上調(diào);ZmBRI1超表達(dá)株系line5對ABA的敏感性最低,物質(zhì)代謝相對穩(wěn)定,脅迫蛋白上調(diào)的倍數(shù)較小,緩解了 ABA對種子萌發(fā)和幼苗生長的抑制作用�；プ骶W(wǎng)絡(luò)分析表明,BR信號主要是通過BIN2與ABI5進行互作,影響ABA信號途徑的抗氧化酶系統(tǒng)、鈣離子信號、MAPK激酶途徑、以及ZIP轉(zhuǎn)錄因子等,從而抑制ABA信號下游基因的表達(dá),促進種子萌發(fā)和幼苗生長。
[Abstract]:Brassinolide (BRs) is an important steroid hormone, which plays an important regulatory role in plant growth and development. At the same time, it is also involved in the regulation of plant response to stress. In this paper, the effects of abiotic stress, such as drought, on plant growth and other problems, such as increasing density or improper management of fertilizer in maize production, have led to the decline of population quality. Using BRs to regulate plant type and response to stress, the growth effect and regulation mechanism of maize were regulated by Br inhibitor Pcz, which provided the basis for the application of Pcz in maize plant shape. The Br receptor gene ZmBRI1 of maize was cloned and its function was analyzed. The protein regulation mechanism of Br signal expression in response to ABA was studied. In order to reveal the theoretical basis of the interaction between Br and ABA in response to stress, the main results were as follows: 1. Pcz treatment significantly inhibited the growth of maize mesocotyl and coleoptile, decreased plant height, shortened the length of leaf and leaf sheath, and reduced the angle between leaves. At the same time, the longitudinal elongation of leaf and leaf sheath cells was significantly inhibited, the arrangement of leaf occipital cells was changed from loose to tight, the content of gibberellin (GA) in maize was decreased, the expression of GA3ox was down-regulated, and the expression of GA2ox5 and GA2ox8, which was inactivated by GA, was upregulated. The expression of GA synthase GA20ox1 was up-regulated and then down-regulated, and the content of Br-R was decreased, but the expression of CPD and DWF4 was up-regulated by feedback regulation. The down-regulated expression of exzr4xyltransferase / hydrolase gene XTH1 XET1 and xylglucan intra glycosyltransferase / hydrolase gene XTH1 XET1 was down-regulated. By using homologous cloning method, a rapesinolactone receptor protein encoding gene ZmBRI1 was obtained from maize inbred line B73. The length of ZmBRI1 was 3369 BP, encoding 1122 amino acids. Subcellular localization analysis showed that ZmBRI1 protein was located on the cell membrane and expressed in all tissues and organs of maize, and ZmBRI1 was introduced into Arabidopsis thaliana Br insensitive mutant bri1-5 by transgenic technique to repair the phenotype of the mutant bri1-5, such as plant height. Leaf morphology and pod size, Br signal of mutant bri1-5, such as root length, Hypocotyl and the expression of Br synthase gene CPDN DWF4, were repaired. Under ABA treatment, line2 and line5 increased seed germination rate and root length significantly. The expression of ABA response gene RS29Agna RD29BHABI5 and RAB18 was decreased, and the sensitivity to ABA was weakened. 3) the proteome of wild type Ws, mutant bri1-5 and ZmBRI1 superexpression line line5 were analyzed during seedling growth, and a total of 5479 proteins were identified. The identified differential proteins are related to primary and secondary metabolism, energy and transport, cell growth and division, protein synthesis and degradation, transcription and signal transduction, The mutant bri1-5 was most sensitive to ABA, the balance of substance metabolism, energy transformation and protein synthesis was broken, and the expression of stress response gene up-regulated ZmBRI1 strain line5 had the lowest sensitivity to ABA. Substance metabolism was relatively stable, stress protein upregulation was small, which alleviated the inhibitory effect of ABA on seed germination and seedling growth. The interaction network analysis showed that the signal of Br was mainly interacted by BIN2 and ABI5. The antioxidant enzyme system, Ca ~ (2 +) signaling MAPK kinase pathway and ZIP transcription factors, which affect the ABA signaling pathway, can inhibit the expression of downstream genes of ABA signal and promote seed germination and seedling growth.
【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】：S513

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