本文選題：小麥　切入點(diǎn)：Ta　出處：《西北農(nóng)林科技大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：赤霉素(GA)在種子發(fā)育過程中發(fā)揮著非常重要的作用,植物通過對(duì)GA信號(hào)感知和轉(zhuǎn)導(dǎo)從而調(diào)控種子發(fā)育過程。E3泛素連接酶廣泛參與到GA信號(hào)感知和轉(zhuǎn)導(dǎo),從而調(diào)控GA信號(hào)途徑介導(dǎo)籽粒發(fā)育過程。TaGW2-6A負(fù)調(diào)控籽粒的大小,研究發(fā)現(xiàn)TaGW2-6A編碼RING E3泛素連接酶。本課題前期研究發(fā)現(xiàn)TaGW2-6A編碼區(qū)中第八外顯子977bp處“T”堿基的插入導(dǎo)致小麥的粒寬和粒重顯著增加。本研究利用中國春CS(野生型)及其近等基因系材料NIL31(T堿基插入突變型)研究了GA相關(guān)基因在花后3,6,9,12,15,20,25天的轉(zhuǎn)錄水平和表達(dá)模式。通過外源GA處理和種子萌發(fā)實(shí)驗(yàn)驗(yàn)證TaGW2-6A不同等位變異對(duì)GA的反應(yīng),從而解析TaGW2-6A等位變異促進(jìn)籽粒變大原因。得到以下主要結(jié)果:1.花后3-25天,NIL31中籽粒胚乳鮮重和干重均高于CS。同時(shí)我們發(fā)現(xiàn),9-20天,NIL31中干物質(zhì)積累速率顯著高于CS。表明,TaGW2-6A等位變異通過影響籽粒灌漿及干物質(zhì)積累速率進(jìn)而控制籽粒的大小。2.實(shí)時(shí)定量PCR結(jié)果表明,GA合成基因(GA 3-oxidase)和GA響應(yīng)基因(GASA4)在NIL31具有比CS更高的表達(dá)水平。此外,NIL31中內(nèi)源GA含量水平顯著高于CS。表明,NIL31中高表達(dá)的GA 3-oxidase促進(jìn)GA合成,使得GASA4表達(dá)量升高,進(jìn)而促進(jìn)胚乳細(xì)胞分裂與發(fā)育,從而影響種子的大小。此外,GA合成和響應(yīng)相關(guān)基因與TaGW2-6A表達(dá)模式分析發(fā)現(xiàn),隨著TaGW2-6A表達(dá)量的降低,GA合成和響應(yīng)相關(guān)基因表達(dá)量逐漸升高,表明TaGW2-6A與GA相關(guān)基因具有負(fù)相關(guān)的關(guān)系。3.外源GA處理實(shí)驗(yàn)結(jié)果發(fā)現(xiàn),CS中,和未處理相比,胚乳細(xì)胞增大,數(shù)目增多,最終籽粒變大。此外,GA 3-oxidase和GASA4基因表達(dá)水平升高,內(nèi)源GA含量水平升高。表明,外源GA處理促進(jìn)了CS中GA 3-oxidase基因的表達(dá),促進(jìn)內(nèi)源GA含量增加,使得GASA4表達(dá)量升高,進(jìn)而影響胚乳細(xì)胞的發(fā)育最終導(dǎo)致籽粒變大。然而,在NIL31中,和未處理相比,胚乳細(xì)胞變小、數(shù)目減少、最終籽粒變小、內(nèi)源GA含量水平降低。此外,GA 3-oxidase水平顯著升高,但GASA4表達(dá)水平降低。由于GASA4基因的低表達(dá),進(jìn)而影響胚乳細(xì)胞的發(fā)育最終導(dǎo)致籽粒的變小。其原因是NIL31本身就有高含量的內(nèi)源GA,當(dāng)受到外源GA刺激時(shí),GA響應(yīng)蛋白作為GA信號(hào)轉(zhuǎn)導(dǎo)的分子開關(guān)從而負(fù)調(diào)節(jié)GA信號(hào),GA 3-oxidase受到負(fù)反饋調(diào)節(jié),從而調(diào)控植物體內(nèi)GA平衡,使GASA4低水平表達(dá),從而引起籽粒的變化。4.種子萌發(fā)實(shí)驗(yàn)結(jié)果表明,GA處理前,NIL31具有比CS高的萌發(fā)率和發(fā)芽勢(shì),此外,NIL31莖的生長速度也明顯高于CS。此外,NIL31籽粒中內(nèi)源GA含量水平顯著高于CS。表明,NIL31中GA合成基因、響應(yīng)基因較高表達(dá)水平,使其體內(nèi)含有較高水平的GA,進(jìn)而促進(jìn)種子萌發(fā)和植物整個(gè)生長過程。GA處理后,種子萌發(fā)實(shí)驗(yàn)發(fā)現(xiàn),CS和NIL31表現(xiàn)出截然相反的萌發(fā)效果�？梢�,植物體內(nèi)GA水平較高時(shí),外源GA的使用,由于負(fù)反饋調(diào)節(jié)反而抑制GASA4的表達(dá),進(jìn)而影響了種子發(fā)育。我們推測(cè),TaGW2-6A與GA信號(hào)協(xié)同作用來調(diào)節(jié)小麥籽粒的發(fā)育。
[Abstract]:Gibberellin (GA) plays a very important role in the process of seed development in plants, the GA signal perception and transduction to regulate seed development process of.E3 ubiquitin ligase GA is widely involved in signal perception and transduction by regulating GA signaling pathway mediated negative regulation of grain development process of.TaGW2-6A grain size, found that TaGW2-6A encoding RING E3 ubiquitin ligase. Our previous study found that eighth of the TaGW2-6A encoding region of exon 977bp of "T" base insertion resulted in wheat grain width and grain weight increased significantly. This study uses Chinese spring CS (wild type) and its near isogenic lines (NIL31 T insertion mutant) of GA gene in the days after flowering 3,6,9,12,15,20,25 transcription and expression pattern. Germination response experimental verification TaGW2-6A different alleles of GA by exogenous GA and TaGW2-6A seeds, and analysis A variation to promote larger grain. The main results are as follows: 3-25 1. days after anthesis, NIL31 in grain endosperm of fresh weight and dry weight were significantly higher than that of CS. at the same time, we found that 9-20 days, dry matter accumulation rate of NIL31 was significantly higher than that of CS. showed that the size of.2. real-time quantitative PCR results TaGW2-6A alleles by grain filling and influence the rate of dry matter accumulation and grain control showed that the synthesis of GA gene (GA 3-oxidase) and GA (GASA4) with responsive gene expression level higher than CS in NIL31. In addition, the level of endogenous GA content in NIL31 was significantly higher than that of CS., high expression of NIL31 in GA 3-oxidase to promote GA synthesis, the GASA4 expression increased, and then promote the endosperm cell division and development, thus affecting seed size. In addition, GA synthesis and response related genes and TaGW2-6A expression pattern analysis showed that the expression of TaGW2-6A with decreased GA synthesis and response related genes Because the expression level increased gradually, showed that TaGW2-6A and GA related genes have a negative correlation between the.3. results showed that exogenous GA treatment, CS, compared with the untreated, the endosperm cells increased, the increase of the number of final grain size. In addition, increased the expression level of GA 3-oxidase and GASA4 gene, increased the content of endogenous GA level shows. That exogenous GA treatment promoted the expression of CS in GA of 3-oxidase gene, promote endogenous GA content increased, the expression of GASA4 increased, thereby affecting endosperm cell development leads to larger grain. However, in NIL31, and the untreated than endosperm cells become smaller, the number of final grain becomes small, endogenous the content of GA decreased. In addition, GA 3-oxidase increased significantly, but the expression level of GASA4 decreased. Due to the low expression of GASA4 gene, thereby affecting endosperm cell development resulting in grain becomes smaller. The reason is that NIL31 itself High content of endogenous GA, when subjected to exogenous GA stimulation, GA response protein as a molecular switch and signal transduction of GA negatively regulates GA signaling, GA 3-oxidase by negative feedback regulation, and regulation of plant GA balance, the low expression of GASA4, the experiment result showed that the changes of.4. seeds germination, before GA treatment NIL31 is higher than CS, the germination rate and germination potential, in addition, the growth rate of NIL31 stem was significantly higher than CS. in addition, the level of endogenous GA content in grains was higher than that of CS. NIL31 showed that the GA synthetic gene NIL31, gene expression in response to the higher level, the body contains higher levels of GA, and then promote the seed germination and the whole plant growth process after.GA treatment, the seed germination experiment showed that CS and NIL31 showed the opposite germination effect. Therefore, the level of GA in higher plants, the use of exogenous GA, but due to the negative feedback regulation Inhibiting the expression of GASA4 and further affecting the development of seeds, we speculate that TaGW2-6A and GA signals are synergistic to regulate the development of wheat grain.

【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S512.1

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