棉花茉莉酸信号基因NINJA和DFR功能鉴定
发布时间：2018-07-31 05:47
【摘要】：植物在抵抗生物和非生物胁迫时,往往限制其生长。尽管在自然环境与农业生态系统中生长与防御的平衡对植物的产量起到重要影响,且茉莉酸(jasmonic acid,JA)信号途径被认为在调节这个平衡过程中起到重要作用,但是对调节植物生长与防御的分子机制的了解非常贫乏。本研究分析了受JA信号途径影响的棉花两个基因的功能。首先,从陆地棉发育早期的纤维中克隆得到一个DFR同源基因,编码355个氨基酸,命名为GhDFR1。通过病毒介导的基因沉默技术发现在棉花中沉默GhDFR1能够显著的减少花青素、原花青素的积累。基于LC-MS分析,GhDFR1沉默植株相比对照植株纤维中两个原花青素单体,( )-epicatachin表儿茶素和( )-epigallocatachin表没食子儿茶素的含量显著的下降,却出现两种新的单体( )-catachin儿茶素和( )-gallocatachin没食子儿茶素。GhDFR1在拟南芥突变体TT3中异源表达能够重建花青素生物合成途径,回复原花青素在拟南芥种皮中积累。同时,本研究分离获得一个棉花NINJA(novel interactor of JA ZIM-domain)基因,其包含一个1305 bp的开放阅读框,将其命名为GhNINJA。GhNINJA基因编码434个氨基酸。根据qRT-PCR分析,GhNINJA在根中优势表达,且表达水平受到大丽轮枝菌侵染诱导。通过病毒介导的基因沉默技术,获得GhNINJA棉花沉默植株,靶基因表达水平仅相当于对照的6%。表型观察发现沉默植株侧根生长受到显著抑制。通过显微观察分析,结果表明GhNINJA沉默植株根组织分化区细胞伸长受到显著抑制,其细胞长度显著短于对照。同时,接种大丽轮枝菌分析,结果表明沉默植株的抗病性显著提高,进一步抗病标记基因分析表明,沉默植株中抗病标记基因PDF1.2和PR4均上调表达,可能造成沉默植株抗病性提高。总之,GhDFR1是棉花花青素、原花青素生物合成途径的重要功能基因,决定花青素和原花青素的积累水平和类型,也是JA信号途径调控的下游靶基因。GhNINJA是棉花JA信号途径中一个重要的抑制子,通过植物间激素的信号网络不仅调控侧根的生长,而且在植物抗病响应中起着重要作用。这两个基因的功能分析为棉花生长和抗性之间的调控关系研究提供了基础数据,也为选育优良的抗逆基因以及相关性状品种的植物改良和分子育种提科学依据。
[Abstract]:Plants tend to limit their growth when they resist biotic and abiotic stresses. Although the balance between growth and defense in the natural environment and agro-ecosystem plays an important role in plant yield, the jasmonic acido JA signaling pathway is thought to play an important role in regulating this balance. But little is known about the molecular mechanisms that regulate plant growth and defense. This study analyzed the function of two genes in cotton affected by JA signaling pathway. Firstly, a homologous gene of DFR was cloned from the early fiber of Upland cotton, encoding 355 amino acids and named GhDFR1. Viral mediated gene silencing technique showed that silencing GhDFR1 in cotton could significantly reduce the accumulation of anthocyanins and procyanidins. The contents of catechin, epicatachin and epigallocatachin in two proanthocyanidins monomers in the fiber of control plants were significantly decreased based on LC-MS analysis. However, two new monomers, catachin catechin and catechin, were expressed in Arabidopsis mutants TT3. The heterologous expression of catechin. GhDFR1 could reconstruct the biosynthesis pathway of anthocyanin and restore the accumulation of proanthocyanidins in the seed coat of Arabidopsis thaliana. At the same time, a cotton NINJA (novel interactor of JA ZIM-domain) gene was isolated, which contained a 1305 BP open reading frame and was named GhNINJA.GhNINJA gene encoding 434 amino acids. According to qRT-PCR analysis, GhNINJA was expressed preferentially in root, and the expression level was induced by Rhizoctonia dahliensis. GhNINJA cotton silencing plants were obtained by virus-mediated gene silencing. The target gene expression level was only 6% of that of the control. Phenotypic observation showed that the lateral root growth of silencing plants was significantly inhibited. The results of microscopic observation and analysis showed that the cell elongation in the root tissue differentiation zone of GhNINJA silencing plants was significantly inhibited and the cell length was significantly shorter than that in the control group. At the same time, the results showed that the resistance of the silencing plants was significantly improved, and the further analysis of the resistance marker genes showed that the expression of PDF1.2 and PR4 were up-regulated in the silencing plants. The disease resistance of silencing plants may be increased. GhDFR1 is an important functional gene of anthocyanin and procyanidin biosynthesis pathway in cotton, which determines the accumulation level and type of anthocyanins and procyanidins. GhNINJA, a downstream target gene regulated by JA signaling pathway, is an important inhibitor of JA signaling pathway in cotton. It not only regulates the growth of lateral roots but also plays an important role in plant disease resistance response through interplant hormone signaling network. The functional analysis of the two genes provides basic data for the study of the regulatory relationship between cotton growth and resistance, and also provides scientific basis for the selection of excellent stress resistance genes and for plant improvement and molecular breeding of related traits.
【学位授予单位】：吉首大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S562
本文编号：2154617
[Abstract]:Plants tend to limit their growth when they resist biotic and abiotic stresses. Although the balance between growth and defense in the natural environment and agro-ecosystem plays an important role in plant yield, the jasmonic acido JA signaling pathway is thought to play an important role in regulating this balance. But little is known about the molecular mechanisms that regulate plant growth and defense. This study analyzed the function of two genes in cotton affected by JA signaling pathway. Firstly, a homologous gene of DFR was cloned from the early fiber of Upland cotton, encoding 355 amino acids and named GhDFR1. Viral mediated gene silencing technique showed that silencing GhDFR1 in cotton could significantly reduce the accumulation of anthocyanins and procyanidins. The contents of catechin, epicatachin and epigallocatachin in two proanthocyanidins monomers in the fiber of control plants were significantly decreased based on LC-MS analysis. However, two new monomers, catachin catechin and catechin, were expressed in Arabidopsis mutants TT3. The heterologous expression of catechin. GhDFR1 could reconstruct the biosynthesis pathway of anthocyanin and restore the accumulation of proanthocyanidins in the seed coat of Arabidopsis thaliana. At the same time, a cotton NINJA (novel interactor of JA ZIM-domain) gene was isolated, which contained a 1305 BP open reading frame and was named GhNINJA.GhNINJA gene encoding 434 amino acids. According to qRT-PCR analysis, GhNINJA was expressed preferentially in root, and the expression level was induced by Rhizoctonia dahliensis. GhNINJA cotton silencing plants were obtained by virus-mediated gene silencing. The target gene expression level was only 6% of that of the control. Phenotypic observation showed that the lateral root growth of silencing plants was significantly inhibited. The results of microscopic observation and analysis showed that the cell elongation in the root tissue differentiation zone of GhNINJA silencing plants was significantly inhibited and the cell length was significantly shorter than that in the control group. At the same time, the results showed that the resistance of the silencing plants was significantly improved, and the further analysis of the resistance marker genes showed that the expression of PDF1.2 and PR4 were up-regulated in the silencing plants. The disease resistance of silencing plants may be increased. GhDFR1 is an important functional gene of anthocyanin and procyanidin biosynthesis pathway in cotton, which determines the accumulation level and type of anthocyanins and procyanidins. GhNINJA, a downstream target gene regulated by JA signaling pathway, is an important inhibitor of JA signaling pathway in cotton. It not only regulates the growth of lateral roots but also plays an important role in plant disease resistance response through interplant hormone signaling network. The functional analysis of the two genes provides basic data for the study of the regulatory relationship between cotton growth and resistance, and also provides scientific basis for the selection of excellent stress resistance genes and for plant improvement and molecular breeding of related traits.
【学位授予单位】：吉首大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S562
【参考文献】
相关期刊论文 前1条
1 马惠;赵鸣;夏晓明;王红艳;董合忠;;Bt棉对棉叶螨发生的影响及与次生代谢物质的关系[J];棉花学报;2012年06期
，本文编号：2154617
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