本文選題：水稻　切入點：早衰　出處：《中國農(nóng)業(yè)科學(xué)院》2016年碩士論文

【摘要】：葉片是植物重要的源器官,水稻葉片早衰將導(dǎo)致葉綠素、蛋白質(zhì)等大分子的降解、光合能力降低,并伴隨活性氧的積累、衰老相關(guān)基因表達(dá)發(fā)生變化、細(xì)胞死亡等現(xiàn)象,嚴(yán)重影響稻米產(chǎn)量及品質(zhì)。因此深入研究早衰形成的分子機理對控制和延緩衰老的發(fā)生具有重要的意義。本研究通過對早衰突變體psls1(premature senescence leaf with spots)形態(tài)、生理等多方面分析,并結(jié)合圖位克隆的方法最終確定候選基因PSLS1:一個編碼鐵氧還依賴的谷氨酸合酶(Fd-GOGAT)基因的突變導(dǎo)致了早衰表型的發(fā)生。主要研究結(jié)果如下:1.突變體psls1幼苗期生長正常,發(fā)育到7葉期以后,葉片自下而上逐漸黃化褪綠至枯萎(黃化由葉尖開始向內(nèi)蔓延并伴有黃褐色小斑點);生長后期突變體的衰老明顯加快,植株整體呈黃褐色枯死狀態(tài)。在成熟期,與野生型相比,psls1株高、分蘗數(shù)、主穗長、穗粒數(shù)和千粒重都顯著下降。2.與野生型相比,psls1衰老葉片葉綠素含量顯著下降,過氧化氫積累、細(xì)胞死亡增加,透射電鏡顯示psls1葉綠體膜界面不清晰,類囊體基粒片層模糊,嗜餓顆粒明顯增多。3.遺傳分析表明,psls1受一對隱性基因控制,利用psls1×IRAT129雜交組合F2分離群體中的1690個早衰個體,將目的基因定位在第7染色體長臂末端分子標(biāo)記ZS-3和ZS-8之間物理距離89Kb的范圍內(nèi),測序研究發(fā)現(xiàn),區(qū)間內(nèi)一個編碼鐵氧還依賴的谷氨酸合酶(Fd-GOGAT)基因LOC_Os07g46460的第二外顯子末位的G被替換為A,導(dǎo)致轉(zhuǎn)錄本的錯誤剪切,相對于野生型,突變體的cDNA缺失了57bp堿基片段,因此,初步認(rèn)定LOC_Os07g46460為可能的候選基因,暫命名為PSLS1。4.同前人報道相一致,PSLS1在水稻根、莖、葉、葉鞘、穗部均有表達(dá),且在葉片中表達(dá)量最高;PSLS1基因的表達(dá)水平在野生型與psls1幼苗中無差異,待分蘗期早衰表型出現(xiàn)后,psls1中PSLS1表達(dá)量顯著下降,對谷氨酸酶活性測定結(jié)果也表明psls1發(fā)生早衰后谷氨酸酶活性顯著降低。5.psls1抽穗期上部葉片中游離氨基酸的含量與野生型存在顯著差異,Glu、Ser、Gly、Ala、Val、Ile的含量顯著下降,而Asn、PSer、Thr、Cys、Phe、His、Orn、Lys、Arg的含量卻顯著增加,表明在psls1突變體葉片中氨基酸代謝紊亂。在水培條件下對野生型和psls1進(jìn)行不同濃度氮素外施處理,發(fā)現(xiàn)psls1對低氮條件響應(yīng)。表明突變體的早衰可能是由于PSLS1的突變使得谷氨酸合酶失活,突變體氮代謝異常引起的。
[Abstract]:Leaf is an important source organ of plants. Premature senescence of rice leaves will lead to the degradation of chlorophyll, protein and other macromolecules, the decrease of photosynthetic ability, the accumulation of reactive oxygen species, the change of senescence related gene expression and cell death, etc.The yield and quality of rice are seriously affected.Therefore, it is of great significance to study the molecular mechanism of premature senescence to control and delay the occurrence of senescence.In this study, morphology and physiology of psls1(premature senescence leaf with spots were analyzed.The mutation of the candidate gene PSLS1: a glutamate synthase (Fd-GOGATT) gene that encodes ferric oxide dependence resulted in the occurrence of premature senescence phenotype.The main results are as follows: 1.The mutant psls1 grew normally at seedling stage, and after developing to 7 leaf stage, the leaves gradually yellowed and faded to wither from top to top (the yellowish brown blotches spread inwards from the tip of the leaf, and the senescence of the mutant accelerated obviously in the later stage of growth.The whole plant was brown and dead.In the mature stage, the plant height, tiller number, main ear length, grain number per ear and 1000-grain weight of psls 1 decreased significantly compared with wild type.Compared with the wild type, the chlorophyll content, hydrogen peroxide accumulation and cell death in senescent leaves of psls1 decreased significantly. Transmission electron microscopy showed that the interface of chloroplast membrane of psls1 was not clear, the lamellae of thylakoid granulocyte was blurred, and the number of eosinophilic granules increased significantly.Genetic analysis showed that psls1 was controlled by a pair of recessive genes. The target gene was located within the range of the physical distance 89Kb between ZS-3 and ZS-8 in the long arm end of chromosome 7 by using 1690 premature senility individuals in F _ 2 segregated population of psls1 脳 IRAT129 hybrid.Sequencing results showed that G at the end of the second exon of a feroxide-dependent glutamate synthase (Fd-GOGATT) gene in the region was replaced with A, resulting in miscut of the transcripts and the deletion of the 57bp base fragment in the cDNA of the mutant relative to the wild type.Therefore, LOC_Os07g46460 was identified as a possible candidate gene, tentatively named PSLS 1.4.In accordance with previous reports, PSLS1 was expressed in roots, stems, leaves, sheaths and panicles of rice, and the highest expression level of PSLS1 gene was found in wild type and psls1 seedlings.After the phenotype of premature senescence appeared at tillering stage, the expression of PSLS1 in pslS1 decreased significantly.The results showed that amino acid metabolism was disturbed in the leaves of psls1 mutants.Wild type and psls1 were treated with different concentrations of nitrogen under hydroponic conditions, and the response of psls1 to low nitrogen condition was found.It is suggested that the premature senescence of the mutant may be due to the inactivation of glutamate synthase caused by the mutation of PSLS1 and the abnormal nitrogen metabolism of the mutant.
【學(xué)位授予單位】：中國農(nóng)業(yè)科學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：S511

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 蔣德安,陸慶,翁曉燕,鄭炳松,奚海福;水稻劍葉衰老期Rubisco活化酶對Rubisco活力和光合速率的調(diào)節(jié)(英文)[J];浙江大學(xué)學(xué)報(農(nóng)業(yè)與生命科學(xué)版);2000年02期

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本文編號：1700808