本文關(guān)鍵詞： 蘋果 生長(zhǎng)素 矮化 砧木 短枝型　出處：《西北農(nóng)林科技大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：蘋果矮化密植栽培是世界蘋果發(fā)展的趨勢(shì),實(shí)現(xiàn)蘋果矮化栽培最主要途徑是利用好矮化砧木及短枝型品種。雖然矮化砧木及短枝型品種在生產(chǎn)中普遍應(yīng)用,但矮化機(jī)理,特別是分子機(jī)理尚未完全揭示。本文分析了長(zhǎng)富2號(hào)/M9(矮化砧木)和長(zhǎng)富2號(hào)/MM106(半喬化砧木)生長(zhǎng)素代謝相關(guān)基因的表達(dá)情況,研究了幾個(gè)關(guān)鍵基因在致矮中的功能和作用。利用miRNA組測(cè)序手段,分析了普通型富士長(zhǎng)富2號(hào)和短枝芽變煙富6號(hào)中枝條發(fā)育相關(guān)差異miRNA。從生長(zhǎng)素和miRNA角度,揭示矮化砧木和短枝型品種的致矮機(jī)理,豐富蘋果矮化調(diào)控理論。取得的主要研究結(jié)果如下:1、對(duì)長(zhǎng)富2號(hào)/M9和長(zhǎng)富2號(hào)/MM106葉片、接穗韌皮部、砧木韌皮部和根系中的生長(zhǎng)素含量、生長(zhǎng)素合成基因、生長(zhǎng)素運(yùn)輸基因和生長(zhǎng)素代謝相關(guān)基因進(jìn)行了分析,結(jié)果發(fā)現(xiàn),長(zhǎng)富2號(hào)/M9中生長(zhǎng)素的濃度低于長(zhǎng)富2號(hào)/MM106。長(zhǎng)富2號(hào)/M9砧木韌皮部的生長(zhǎng)素濃度高于其接穗。在長(zhǎng)富2號(hào)/M9葉片和根系中,生長(zhǎng)素合成基因MdYUCCA8和MdYUCCA10a表達(dá)顯著低于長(zhǎng)富2號(hào)/MM106。長(zhǎng)富2號(hào)/M9砧木韌皮部和根系中,生長(zhǎng)素運(yùn)輸基因MdPIN1b和Md PIN8a顯著低于長(zhǎng)富2號(hào)/MM106。長(zhǎng)富2號(hào)/M9中的生長(zhǎng)素軛合酶基因MdGH3-5b和MdGH3-9a顯著低于長(zhǎng)富2號(hào)/MM106。然而,長(zhǎng)富2號(hào)/M9中的生長(zhǎng)素水解酶基因MdIAR3c和MdILL6c顯著高于長(zhǎng)富2號(hào)/MM106。2、在M9和MM106中克隆了MdYUCCA8基因,該基因編碼1272 bp核苷酸,二者序列相似性99.9%,編碼423個(gè)氨基酸�？寺×薓dYUCCA8編碼區(qū)上游1600 bp啟動(dòng)子序列,二者相似性為99.6%。MdYUCCA8啟動(dòng)子受到GA、MJ和Br誘導(dǎo),亞精胺和乙烯利顯著抑制了MdYUCCA8啟動(dòng)子活性。高溫促進(jìn)MdYUCCA8的表達(dá),煙草葉片雙熒光素酶瞬時(shí)表達(dá)實(shí)驗(yàn)證明MdPIF4a與MdYUCCA8啟動(dòng)子互作。在擬南芥中異位表達(dá)MdYUCCA8,促進(jìn)了擬南芥植株花莖的生長(zhǎng)、頂端優(yōu)勢(shì)顯著增加、側(cè)枝減少、角果畸形、促進(jìn)了根系的生長(zhǎng),側(cè)根數(shù)量和側(cè)根長(zhǎng)度顯著高于野生型。3、在M9和MM106中克隆了MdYUCCA10a基因,該基因編碼1149 bp核苷酸,二者序列相似性99.9%,編碼382個(gè)氨基酸�？寺×薓dYUCCA10a編碼區(qū)上游1800 bp啟動(dòng)子序列,二者相似性為99.7%。MdYUCCA10a啟動(dòng)子受到ABA、Spe誘導(dǎo),乙烯利顯著抑制MdYUCCA10a啟動(dòng)子活性。在擬南芥中異位表達(dá)MdYUCCA10a,促進(jìn)了擬南芥植株花莖的生長(zhǎng),促進(jìn)了根系的生長(zhǎng),主根長(zhǎng)度、側(cè)根數(shù)量和側(cè)根長(zhǎng)度顯著高于野生型。4、在M9和MM106中克隆了MdABCB19基因,其編碼1250個(gè)氨基酸,含有兩個(gè)ABC_membrance結(jié)構(gòu)域,兩個(gè)ABC_tran結(jié)構(gòu)域。該序列在M9和MM106中存在一個(gè)非同義SNP,編碼氨基酸由A突變?yōu)镾,導(dǎo)致M9α螺旋少了一段。MdABCB19在砧木M9和MM106、長(zhǎng)富2號(hào)/M9和長(zhǎng)富2號(hào)/MM106均差異表達(dá)。啟動(dòng)子序列分析發(fā)現(xiàn)M9的MdABCB19啟動(dòng)子在起始密碼子上游170 bp處有“CTCTGT”6個(gè)堿基缺失,導(dǎo)致M9缺失了一個(gè)5’UTR Py-rich stretch motif。MM106 MdABCB19的啟動(dòng)子活性高于M9 MdABCB19,并且MdABCB19啟動(dòng)子受光照調(diào)控。MdABCB19啟動(dòng)子活性隨著5’UTR Py-rich stretch motif元件數(shù)目的增加而增強(qiáng)。5、在長(zhǎng)富2號(hào)及短枝型芽變煙富6號(hào)頂梢中,共發(fā)現(xiàn)700個(gè)成熟的miRNA,包括202個(gè)蘋果已知mi RNA和498新miRNA。長(zhǎng)富2號(hào)和煙富6號(hào)中有135個(gè)差異表達(dá)的miRNA,大多數(shù)差異的已知miRNA在煙富6號(hào)中下調(diào)表達(dá)。煙富6號(hào)枝條頂梢IAA、GA、ZR和ABA含量都低于長(zhǎng)富2號(hào)。噴施GA可以促進(jìn)煙富6號(hào)短枝生長(zhǎng),變?yōu)殚L(zhǎng)枝。煙富6號(hào)頂梢中細(xì)胞伸長(zhǎng)和細(xì)胞分裂相關(guān)基因的表達(dá)水平都顯著低于長(zhǎng)富2號(hào)。綜上所述,生長(zhǎng)素在砧木誘導(dǎo)接穗矮化中起著重要作用,長(zhǎng)富2號(hào)/M9中低表達(dá)的生長(zhǎng)素合成基因MdYUCCA8和MdYUCCA10a可能導(dǎo)致長(zhǎng)富2號(hào)/M9中低的生長(zhǎng)素含量。M9 MdABCB19啟動(dòng)子5’UTR Py-rich stretch motif缺失,可能造成M9中MdABCB19表達(dá)量降低,生長(zhǎng)素運(yùn)輸減弱,植株矮小。GA和miRNA在煙富6號(hào)短枝發(fā)育中起著關(guān)鍵調(diào)控作用。
[Abstract]:The dwarf and close planting apple is the world trend of development, the realization of Apple dwarfing cultivation the main way is to use a good rootstock and spur type. Although the dwarf rootstock and spur type which is widely used in the production, but the Dwarfing mechanism, especially the molecular mechanism has not been fully revealed. This paper analyzes the /M9 (dwarf Nagafu No. 2 stock) and /MM106 (Joe Nagafu No. 2 half of the stock) the expression of auxin metabolism related genes, the study of several key genes in the function and role of dwarfing in group miRNA. The sequencing methods, analyzes the common type of Fuji Nagafu No. 2 and No. 6 short branch bud smoke rich in shoot development related differences from miRNA. auxin and miRNA angle, reveals the Dwarfing mechanism of Dwarfing Rootstock and spur type variety, rich apple dwarf control theory. The main results are as follows: 1, the Nagafu No. 2 /M9 and Nagafu No. 2 leaf /MM106, scion toughness Firstly, the content of IAA in root and rootstock phloem, auxin biosynthesis gene, auxin transport genes and auxin metabolism related genes were analyzed. The results showed that auxin concentration of auxin Nagafu No. 2 in /M9 is lower than that of Nagafu No. 2 /MM106. Nagafu No. 2 /M9 rootstock phloem is higher than that of the scion. In Nagafu No. 2 /M9 leaves and roots, expression of auxin synthesis genes MdYUCCA8 and MdYUCCA10a were significantly lower than the Nagafu No. 2 /MM106. Nagafu No. 2 /M9 rootstock phloem and root, auxin transport genes MdPIN1b and Md were significantly lower than that of PIN8a / MM106. yoke Nagafu No. 2 auxin Nagafu No. 2 in /M9 synthase gene MdGH3-5b and MdGH3-9a were significantly lower than the Nagafu No. 2 /MM106. however. Nagafu No. 2 in /M9 MdIAR3c and MdILL6c were significantly higher than that of auxin hydrolase gene Nagafu No. 2 in /MM106.2, M9 and MM106 in MdYUCCA8 gene was cloned and the gene encoding the 1272 BP core The nucleotide sequence similarity of two, 99.9%, encoding 423 amino acids. The cloning of the MdYUCCA8 encoding region 1600 bp upstream promoter sequence, two similarity to 99.6%.MdYUCCA8 promoter by GA, MJ and Br induced by spermidine and ethephon significantly inhibited the MdYUCCA8 promoter activity. The expression of MdYUCCA8 in high temperature to promote the tobacco the leaves of dual luciferase transient expression experiments demonstrated that MdPIF4a and MdYUCCA8 promoter interactions. Ectopic expression of MdYUCCA8 in Arabidopsis thaliana, promote Arabidopsis stem growth, apical dominance increased significantly, branches reduced, pod deformity, promote root growth, root number and root length were significantly higher than that of the wild type.3, M9 and MM106 in clone the MdYUCCA10a gene, the gene encoding 1149 BP nucleotides, two sequence similarity 99.9%, encoding 382 amino acids. The cloning of the MdYUCCA10a encoding region 1800 bp upstream promoter sequence, two similarity For the 99.7%.MdYUCCA10a promoter by ABA, induced by Spe, ethephon significantly inhibited the activity of MdYUCCA10a promoter. Ectopic expression of MdYUCCA10a in Arabidopsis thaliana, promote Arabidopsis stem growth, promote root growth, root length, root number and root length were significantly higher than those in the wild type.4, M9 and MM106 in MdABCB19 gene was cloned. The encoding 1250 amino acids, containing two ABC_membrance domains and two ABC_tran domains. The sequence has a non synonymous SNP in M9 and MM106, encoding amino acid substitution from A to S, resulting in M9 alpha helix is less of a.MdABCB19 in stock M9 and MM106, the expression of /M9 and Fuji Nagafu No. 2 2 No. /MM106 were different. The promoter sequence analysis indicated that the M9 promoter of MdABCB19 "CTCTGT" 6 nucleotide deletion in the upstream of the start codon 170 BP, resulting in a loss of M9 5 UTR Py-rich stretch motif.MM106 MdAB " The CB19 promoter activity was higher than that of M9 MdABCB19, and MdABCB19 promoter by the light regulation of.MdABCB19 promoter activity increased with 5 UTR Py-rich stretch motif "the number of components and enhanced.5, in Nagafu No. 2 and spur type sports smoke rich 6 tops, 700 were found in mature miRNA, including 202 Apple MI is known RNA and 498 new miRNA. Nagafu No. 2 and No. 6 in smoke rich 135 differential expression of miRNA, miRNA is known most differences in tobacco rich down regulates the expression of 6. 6 branches tops smoke rich IAA, GA, ZR and ABA content was lower than that of Nagafu No. 2. Application of GA can promote smoke into the rich 6 short branches grow into long branches. The expression level of tobacco rich cell elongation and cell division related genes in 6 tops are significantly lower than those of Nagafu No. 2. In summary, auxin induced scion plays an important role in Dwarfing Rootstock, low expression of Nagafu No. 2 /M9 synthase gene M auxin DYUCCA8 and MdYUCCA10a may cause Nagafu No. 2 /M9.M9 low auxin MdABCB19 promoter 5 'UTR Py-rich stretch motif deletion, may cause decreased MdABCB19 expression in M9, auxin transport decreased, plant small.GA and miRNA rich 6 short branch development plays a key role in the smoke.

【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：S661.1

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 羅婭,湯浩茹;梨矮化砧木選育與離體繁殖研究進(jìn)展[J];中國(guó)南方果樹;2004年04期

2 任盈;;梨矮化砧木通過(guò)鑒定[J];農(nóng)村百事通;2007年13期

3 沙守峰;張紹鈴;李俊才;;梨矮化砧木的選育及其應(yīng)用研究進(jìn)展[J];北方園藝;2009年08期

4 蔡忠民;李俊才;王家珍;沙守峰;李宏軍;;梨矮化砧木研究進(jìn)展[J];北方果樹;2013年05期

5 ;為什么矮化砧木能使果樹長(zhǎng)得矮小?[J];植物學(xué)雜志;1974年02期

6 小池洋男;郁香荷;;日本正廣泛使用的矮化砧木[J];北方果樹;1987年02期

7 胡晶晶,胡瑩;優(yōu)良的K系梨矮化砧木[J];山西農(nóng)業(yè);2000年12期

8 及華,張海新;梨矮化砧木選育及離休繁殖技術(shù)研究進(jìn)展[J];河北林業(yè)科技;2002年03期

9 趙玉輝,周懷軍,黎鴻慧;經(jīng)濟(jì)林矮化砧木與鑒定方法研究概況[J];河北林果研究;2002年02期

10 孫艷香,呂增仁,張成合,陳段芬;利用系統(tǒng)聚類法進(jìn)行杏矮化砧木預(yù)選的初步研究[J];河北農(nóng)業(yè)大學(xué)學(xué)報(bào);2002年04期

相關(guān)會(huì)議論文 前10條

1 姜淑苓;賈敬賢;馬力;;梨樹矮化砧木的研究與應(yīng)用進(jìn)展(1)[A];中國(guó)園藝學(xué)會(huì)第十屆會(huì)員代表大會(huì)暨學(xué)術(shù)討論會(huì)論文集[C];2005年

2 劉巖;張玉星;張殿生;喬進(jìn)春;許建鋒;;梨樹矮化砧木病毒病及脫病毒研究概況[A];全國(guó)第四屆梨科研、生產(chǎn)與產(chǎn)業(yè)化學(xué)術(shù)研討會(huì)論文集[C];2005年

3 沙廣利;毛慶滿;黃粵;李凌;邵永春;;蘋果矮化砧苗期性狀的遺傳傾向分析[A];中國(guó)園藝學(xué)會(huì)第九屆學(xué)術(shù)年會(huì)論文集[C];2001年

4 聶書海;;淺談我省矮化蘋果發(fā)展的幾個(gè)問題[A];河北省果樹學(xué)會(huì)第十二屆學(xué)術(shù)年會(huì)論文摘要匯編[C];1991年

5 李榮富;蔣親賢;梁艷榮;梁莉;郭喜平;;不同抗寒矮化砧對(duì)金紅蘋果幼樹生長(zhǎng)發(fā)育的影響[A];中國(guó)園藝學(xué)會(huì)第六屆青年學(xué)術(shù)討論會(huì)論文集[C];2004年

6 王蘇珂;李秀根;楊健;王龍;;西洋梨矮化砧木BA-29的組織培養(yǎng)和快速繁殖技術(shù)研究[A];梨科研與生產(chǎn)進(jìn)展（五）[C];2011年

7 金萬(wàn)梅;毛海亮;張強(qiáng);王小偉;楊廷楨;魏欽平;;SH系蘋果矮化砧木的SSR標(biāo)記鑒定(摘要)[A];第二屆全國(guó)果樹分子生物學(xué)學(xué)術(shù)研討會(huì)論文集[C];2009年

8 李文光;裴學(xué)魁;劉魁英;;蘋果矮化材料予選方法的研究[A];河北省果樹學(xué)會(huì)第七屆、第八屆學(xué)術(shù)年會(huì)論文摘要集[C];1981年

9 羅婭;湯浩茹;王小蓉;甘玲;;梨矮化砧S_2、S_5和PDR_(54)試管苗生根研究[A];全國(guó)第四屆梨科研、生產(chǎn)與產(chǎn)業(yè)化學(xué)術(shù)研討會(huì)論文集[C];2005年

10 楊廷楨;王騫;高敬東;李鯤鵬;韓鳳;田建保;;抗寒、抗旱蘋果矮化砧木選育研究初報(bào)[A];2008園藝學(xué)進(jìn)展（第八輯）——中國(guó)園藝學(xué)會(huì)第八屆青年學(xué)術(shù)討論會(huì)暨現(xiàn)代園藝論壇論文集[C];2008年

相關(guān)重要報(bào)紙文章 前10條

1 李丙智;世界蘋果矮化砧木應(yīng)用現(xiàn)狀(二)[N];河南科技報(bào);2009年

2 李丙智;世界蘋果矮化砧木應(yīng)用現(xiàn)狀(一)[N];河南科技報(bào);2009年

3 都文;我國(guó)蘋果矮化砧木發(fā)展緩慢原因分析[N];瓜果蔬菜報(bào).農(nóng)業(yè)信息周刊;2008年

4 國(guó)家蘋果產(chǎn)業(yè)技術(shù)體系首席專家 韓明玉;我國(guó)矮化蘋果園存在的問題[N];河北科技報(bào);2009年

5 李丙智 韓明玉 張林森 張滿讓 許志強(qiáng);我國(guó)蘋果矮化砧木發(fā)展緩慢的原因[N];陜西科技報(bào);2007年

6 李丙智　韓明玉;我國(guó)蘋果矮化砧木為何發(fā)展緩慢？[N];山東科技報(bào);2010年

7 李丙智 韓明玉 張林森 張滿讓 許志強(qiáng);我國(guó)蘋果矮化砧木為何發(fā)展緩慢？[N];山東科技報(bào);2008年

8 楊宇;我國(guó)首次培育出蘋果種子繁育矮化砧木[N];科技日?qǐng)?bào);2007年

9 通訊員 邵陽(yáng)邋劉磊;蘋果無(wú)融合生殖矮化砧木新品系育種成功[N];科技日?qǐng)?bào);2007年

10 方清軍;果樹矮化的有效途徑[N];北京科技報(bào);2003年

相關(guān)博士學(xué)位論文 前3條

1 馮軼;蘋果矮化砧木M9根系IPT5b基因表達(dá)調(diào)控及其致矮機(jī)理的研究[D];中國(guó)農(nóng)業(yè)大學(xué);2017年

2 安海山;蘋果喬化砧、矮化砧根系生長(zhǎng)特性比較研究[D];中國(guó)農(nóng)業(yè)大學(xué);2017年

3 宋春暉;蘋果IAA代謝幾個(gè)關(guān)鍵基因在矮化砧木與miRNA在短枝品種中致矮作用研究[D];西北農(nóng)林科技大學(xué);2017年

相關(guān)碩士學(xué)位論文 前10條

1 王淼淼;幾種新型蘋果矮化砧木的組培快繁技術(shù)研究[D];河北農(nóng)業(yè)大學(xué);2015年

2 王麗;SH40蘋果矮化砧木扦插繁殖技術(shù)及生根機(jī)理初探[D];河北農(nóng)業(yè)大學(xué);2015年

3 吳學(xué)麗;三種蘋果矮砧扦插生根影響因子的研究[D];西北農(nóng)林科技大學(xué);2015年

4 鄭立偉;蘋果矮化砧木T337中BR相關(guān)基因CBB1、miR492及其靶基因BAK1的克隆與表達(dá)分析[D];西北農(nóng)林科技大學(xué);2016年

5 魏林;陜西蘋果矮化砧木應(yīng)用現(xiàn)狀及區(qū)域化分析[D];西北農(nóng)林科技大學(xué);2016年

6 徐穎;梨矮化砧木預(yù)選指標(biāo)及矮化性狀相關(guān)分子標(biāo)記的研究[D];延邊大學(xué);2010年

7 羅婭;梨矮化砧木組織培養(yǎng)快繁的研究[D];四川農(nóng)業(yè)大學(xué);2004年

8 莊得鳳;梨抗寒矮化砧木的預(yù)選及矮化性狀相關(guān)分子標(biāo)記的研究[D];延邊大學(xué);2008年

9 及華;梨S系矮化砧組培快繁技術(shù)體系研究[D];河北農(nóng)業(yè)大學(xué);2002年

10 樸永虎;延邊小香水梨（Pyrus.ussuriensis Maxim）偶然實(shí)生矮化突變體的評(píng)價(jià)研究[D];延邊大學(xué);2010年

，

本文編號(hào)：1442779