本文關(guān)鍵詞： 中間錦雞兒 MYB 表達(dá)分析 啟動(dòng)子克隆　出處：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：干旱、鹽堿、高溫、低溫和氧化等非生物脅迫是世界范圍內(nèi)作物減產(chǎn)的主要原因。應(yīng)用轉(zhuǎn)錄因子改善植物抗逆性已經(jīng)成為當(dāng)今的研究熱點(diǎn)。MYB類轉(zhuǎn)錄因子是植物中最大的轉(zhuǎn)錄因子家族之一,而R2R3-MYB類是植物MYB基因編碼蛋白中最豐富的類型。R2R3-MYB在初生與次生代謝、生長發(fā)育及對生物與非生物脅迫的應(yīng)答等各個(gè)方面具有重要功能。中間錦雞兒(Csragana intermedia)作為重要的保水、防風(fēng)、固沙類灌木,在高寒沙地被作為主要造林灌木樹種之一。本研究以中間錦雞兒為實(shí)驗(yàn)材料克隆到4個(gè)R2R3-MYB類轉(zhuǎn)錄因子,分別命名為CiMYB102、CiMYB31、CiMYB60和CiMYB185,并對其表達(dá)模式及啟動(dòng)子順式作用元件進(jìn)行了分析。主要結(jié)果如下:1.從干旱轉(zhuǎn)錄組數(shù)據(jù)庫中篩選到4個(gè)R2R3-MYB并對其cDNA和gDNA進(jìn)行了克隆。CiMYB102、CiMYB31、CiMY60與 CiMYB185 的 ORF 長度分別為 1107 bp、969 bp、1038 bp與906 bp,分別編碼369、323、346和302個(gè)氨基酸。CiMYB102 CiMYB31、CiMYYB60與CiMYB185基因gDNA序列長度分別為1396 bp、1724 bp、1485 bp和1592 bp,其中CiMYB102、CiYB31和CiMYB60均包含2個(gè)內(nèi)含子和3個(gè)外顯子,而CiM7B185包含1個(gè)內(nèi)含子和2個(gè)外顯子。2.利用實(shí)時(shí)熒光定量PCR技術(shù)分析了 4個(gè)R2R3-MYB基因不同脅迫下的表達(dá)模式。發(fā)現(xiàn)CiMYB31和CiMYB185受到低溫的誘導(dǎo),CiMYB102受脫水、NaCl、ABA和千旱的誘導(dǎo),在脫水、NaCl、UV-B處理下CiMYB60的表達(dá)量均降低。表明四個(gè)R2R3-MYB可能在中間錦雞兒對非生物肋迫的響應(yīng)過程中起作用。3.利用染色體步移技術(shù)克隆了 4個(gè)基因的啟動(dòng)子序列。CiMYB102、CiMYB31、CiMYB60與CiMYB185的ATG上游序列長度分別為1359 bp、902 bp、1848 bp與908 bp,分析顯示啟動(dòng)子序列中包含一些與光反應(yīng)、組織特異性表達(dá)、激素和非生物脅迫相關(guān)的響應(yīng)元件。4.CiMYB102、CiMYTB60及CiMYB31在不同組織部位表達(dá)存在差異,在根中的表達(dá)量均為最低。CiMYB102在莖中表達(dá)量最高,CiMYB31在葉中表達(dá)量最高,CiMYB60在葉與莖中表達(dá)量均相對較高。5.構(gòu)建 了過表達(dá)載體p35s::CiMYB102-GFP、p35s::CiMYB31-GFP和p35s::CiMYB60-GFP,轉(zhuǎn)化野生型擬南芥并篩選得到純合體株系。
[Abstract]:Drought, salt, high temperature. Abiotic stress, such as low temperature and oxidation, is the main cause of crop reduction in the world. The application of transcription factors to improve plant stress resistance has become a hot topic. MYB transcription factors are the largest transcriptional factors in plants. One of the children. R2R3-MYB is the most abundant type of plant MYB gene encoding protein. R2R3-MYB is primary and secondary metabolism. Growth and development as well as response to biological and abiotic stress have important functions. Csragana intermedia is an important water conservation and windbreak. Sand-fixing shrub was used as one of the main afforestation shrub species in alpine sandy land. In this study, four transcription factors of R2R3-MYB were cloned from Caragana intermedia. They were named CiMYB102, CiMYB31, CiMYB60 and CiMYB185, respectively. The expression pattern and promoter cis-acting elements were analyzed. The main results were as follows:. 1. Four R2R3-MYB were screened from the database of drought transcriptome and their cDNA and gDNA were cloned. CiMYB102. The ORF length of CiMYB31C CiMY60 and CiMYB185 were 1038bp and 906bp, respectively. It encodes 369,323,346 and 302 amino acids. CiMYB102 CiMYB31, respectively. The length of gDNA sequence of CiMYYB60 and CiMYB185 gene was 1396 BP, 1724 BP, 1485 BP and 1592 BP, respectively. Both CiMYB102 and CiYB31 and CiMYB60 contained two introns and three exons. However, CiM7B185 contains one intron and two exons. It is analyzed by real-time fluorescence quantitative PCR. Four R2R3-MYB gene expression patterns under different stress. It was found that CiMYB31 and CiMYB185 were induced by low temperature. CiMYB102 was induced by dehydration and drought. The expression of CiMYB60 decreased under UV-B treatment, which suggested that four R2R3-MYB might play a role in the response of Caragana intermedia to abiotic costal forces. 3. Chromosome step technique was used to detect the effect of R2R3-MYB on the response of Caragana intermedia to abiotic costal forces. Up. The promoter sequence of four genes. CiMYB102. The upstream ATG length of CiMYB31C CiMYB60 and CiMYB185 was 1359bpmc902 BP 1848 BP and 908bp respectively. Analysis showed that the promoter sequence contained some response elements related to light response, tissue specific expression, hormone and abiotic stress. 4. CiMYB102. The expression of CiMYTB60 and CiMYB31 were different in different tissues, and the lowest expression was in root. CiMYB102 was the highest in stem. The expression of CiMYB60 in leaves and stems was higher than that in leaves. 5. the overexpression vector p35s:: CiMYB102-GFP was constructed. P35: s: CiMYB31-GFP and p35s: CiMYB60-GFP.The wild type Arabidopsis thaliana was transformed and homozygous lines were screened.
【學(xué)位授予單位】：內(nèi)蒙古農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：Q943.2

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