杨树PtPIF和PtPIN1s基因功能的初步分析
发布时间：2019-05-17 10:22
【摘要】：光敏色素相互作用因子(Phytochrome Interacting Factor,PIF)是碱性螺旋-环-螺旋(basic Helix-Loop-Helix,bHLH)转录因子家族的一员,参与了植物光信号传导。拟南芥基因组共编码7个PIFs,在种子萌发、下胚轴伸长、植物蔽荫反应和叶片衰老等过程具有重要功能,通过调控生长素的合成与分布是其影响植物生长发育的重要途径。木本植物具有不同于草本植物的生长发育过程,因此对光信号的应答也存在差异。研究杨树中PIF基因功能有助于揭示树木光信号响应机制。本研究分析了杨树Pt PIF基因家族成员的系统进化关系、基因结构、保守结构域及不同组织和不同胁迫条件下的表达模式。生长素运输载体PIN是植物特有的,细胞膜极性定位的PIN蛋白可能受到PIF调控生长素的运输改变生长素浓度梯度,从而影响植物生长发育。毛果杨共有15个Pt PIN基因,相对于拟南芥发生了显著扩张。为了探究PtPIN1s在表达及功能上的差异,通过GUS染色分析了PtPIN1s的组织表达差异,并分析了PtPIN1s在不同组织和不同胁迫条件下的表达模式。研究结果如下:1.筛选得到10个PtPIF基因家族成员,是拟南芥7个PIF基因家族成员的1.43倍;系统进化分析发现了4个同源基因对(PtPIF3a/PtPIF3b、PtPIF4/5a/PtPIF4/5b、PtPIF8a/PtPIF8b和PtPIF9a/PtPIF9b);PtPIFs的基因结构和蛋白结构域相对保守;且预测PtPIFs基因启动子区包含激素等胁迫响应元件。表明PtPIF能够响应环境因子的变化,比如光。2.PtPIFs基因具有组织表达特异性并能响应多种非生物胁迫。Pt PIF1、PtPIF8a和PtPIF8b主要在成熟叶中表达,PtPIF3a和Pt PIF9b主要在幼叶中表达;低温处理下,PtPIF1、PtPIF4/5a、PtPIF8b和PtPIF9b的表达量下调,PtPIF10上调;干旱处理下,PtPIF3b和PtPIF4/5b的表达量下调,PtPIF8a上调;盐处理下,PtPIF4/5a的表达量下调,PtPIF3b和PtPIF9a上调。表明PtPIFs能够响应不同的环境胁迫。3.为了验证PtPIFs能够与G-box结合,选取PtPIF3a构建至pGADT7-Rec2载体上,将G-box序列与突变了的G-box序列以三次重复方式串联并克隆至pHIS2载体上。酵母单杂交实验结果表明,PtPIF3a能够与G-box结合,而不能与突变了的G-box结合。Pt PIF3a的结合能力表明PtPIFs有可能通过G-box调控PIN。4.根据系统进化分析的结果将PtPIN1s分为两类,一类是PtPIN1a和PtPIN1b,另一类是PtPIN1c和PtPIN1d。GUS染色结果发现,PtPIN1a和PtPIN1b在不定根根尖处的表达强于PtPIN1c和PtPIN1d,这种表达差异产生于不定根形成的愈伤组织阶段,且不受外源生长素影响。在低温、干旱及盐胁迫下,PtPIN1a/Pt PIN1b和Pt PIN1c/PtPIN1d的表达有不同也有相似之处。低温处理下,PtPIN1b的表达量下调,PtPIN1c上调;干旱处理下PtPIN1b和PtPIN1c的表达量上调,PtPIN1d下调;盐处理下,PtPIN1d的表达量下调。PtPIN1s在响应胁迫上的不同,表明木本植物PIN蛋白在进化和功能上的多样化。本研究预测了杨树PtPIF基因家族成员,并对各成员的基因结构、保守结构域进行了系统分析;检测了PtPIF成员在不同组织及低温、干旱和NaCl胁迫下的表达模式。同时,证明了PtPIN1s在不定根形成及不同非生物胁迫下的不同表达模式。PtPIF3a与G-box的结合能力表明PtPIFs有可能与PIN发生相互作用。研究结果为深入探究杨树PtPIF和PtPIN1s在不同环境下调控植物生长发育的作用机制奠定了基础。
[Abstract]:The phytochrome interaction factor (PIF) is a member of the basic helix-loop-Helix (bHLH) transcription factor family and is involved in the transmission of plant optical signals. The Arabidopsis genome has seven PIFs, and has important function in the process of seed germination, hypocotyl elongation, plant shade reaction and leaf senescence, and the synthesis and distribution of the auxin is an important way to influence the growth and development of the plants. The woody plant has a different growth and development process than the herbaceous plant, so there is a difference in the response of the light signal. The study of PIF gene function in poplar can help to reveal the response mechanism of tree light signal. The system evolution relation, gene structure, conserved domain and expression pattern of different tissues and different stress conditions of the family members of the PIF gene family of poplar were analyzed in this study. The PIN of the auxin transport carrier is unique to the plant, and the PIN protein of the cell membrane polarity can be influenced by the transportation of the PIF regulatory auxin to change the auxin concentration gradient, thereby affecting the growth and development of the plant. A total of 15 Pt PIN genes were found in Populus tomentosa, and a significant expansion was observed in Arabidopsis. In order to study the difference between the expression and function of PtIN1s, the expression of PtPIN1s was analyzed by GUS staining, and the expression pattern of PtIN1s in different tissues and under different stress conditions was analyzed. The results of the study are as follows:1. The gene structure and protein domain of PtPIF3a/ PtPIF3b, PtPIF4/ 5a/ PtPIF4/ 5b, PtPIF8a/ PtPIF8b and PtPIF9a/ PtPIF9b were found by the system evolution analysis, and the gene structure and the protein domain of PtPIFs were relatively conserved. And the predicted PtPIFs gene promoter region comprises a stress response element such as a hormone. It is shown that PtPIF can respond to changes of environmental factors, such as light.2. PtPIFs gene has tissue expression specificity and can respond to various abiotic stress. The expression of PtPIF1, PtPIF8a and PtPIF8b is mainly expressed in the mature leaves, and the expression of PtPIF1, PtPIF4/ 5a, PtPIF8b and PtPIF9b is down-regulated under the low temperature treatment, and the PtPIF10 is up-regulated; in the drought treatment, the expression amount of the PtPIF3b and the PtPIF4/ 5b is down-regulated, and the PtPIF8a is up-regulated; and under the salt treatment, the expression amount of the PtPIF4/ 5a is reduced, PtPIF3b and PtPIF9a are up-regulated. It is shown that PtPIFs can respond to different environmental stresses. To verify that PtPIFs can be combined with G-box, PtPIF3a is selected to be constructed on pGADT7-Rec2 vector, and the G-box sequence and the mutated G-box sequence are connected in series and cloned to the pHIS2 vector in triplicate. The results of single-hybridization experiments show that PtPIF3a can be combined with G-box and can not be combined with the mutated G-box. The binding capacity of Pt PIF3a indicates that PtPIFs has the potential to control the PIN through G-box. According to the results of the system evolution analysis, PtPIN1s were divided into two groups, one is PtPIN1a and PtPIN1b, and the other is PtPIN1c and PtPIN1d. The results of GUS staining show that the expression of PtPIN1a and PtPIN1b at the root tip of the adventitious root is stronger than that of PtPIN1c and PtPIN1d, and the expression difference is generated in the callus stage formed by the adventitious root. And is not affected by exogenous auxin. At low temperature, drought and salt stress, the expression of PtPIN1a/ Pt PIN1b and Pt PIN1c/ PtPIN1d is also similar. Under the low temperature treatment, the expression of PtPIN1b was down-regulated, and the expression of PtPIN1b and PtPIN1c was up-regulated under the drought treatment, and the expression of PtPIN1d was down-regulated under the treatment of salt. The difference of PtPIN1s in response to stress indicates that the PIN protein of the woody plant is diverse in evolution and function. The gene structure and conserved domain of PtPIF gene of poplar were systematically analyzed in this study, and the expression patterns of PtPIF members under different tissues and at low temperature, drought and NaCl stress were detected. At the same time, the different expression patterns of PtIN1s in the formation of adventitious roots and different abiotic stresses are demonstrated. The binding capacity of PtPIF3a with G-box indicates that PtPIFs may interact with the PIN. The results of the study provide a basis for the in-depth study of the mechanism of PtPIF and PtIN1s in the control of plant growth and development in different environments.
【学位授予单位】：中国林业科学研究院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;S792.11
本文编号：2479025
[Abstract]:The phytochrome interaction factor (PIF) is a member of the basic helix-loop-Helix (bHLH) transcription factor family and is involved in the transmission of plant optical signals. The Arabidopsis genome has seven PIFs, and has important function in the process of seed germination, hypocotyl elongation, plant shade reaction and leaf senescence, and the synthesis and distribution of the auxin is an important way to influence the growth and development of the plants. The woody plant has a different growth and development process than the herbaceous plant, so there is a difference in the response of the light signal. The study of PIF gene function in poplar can help to reveal the response mechanism of tree light signal. The system evolution relation, gene structure, conserved domain and expression pattern of different tissues and different stress conditions of the family members of the PIF gene family of poplar were analyzed in this study. The PIN of the auxin transport carrier is unique to the plant, and the PIN protein of the cell membrane polarity can be influenced by the transportation of the PIF regulatory auxin to change the auxin concentration gradient, thereby affecting the growth and development of the plant. A total of 15 Pt PIN genes were found in Populus tomentosa, and a significant expansion was observed in Arabidopsis. In order to study the difference between the expression and function of PtIN1s, the expression of PtPIN1s was analyzed by GUS staining, and the expression pattern of PtIN1s in different tissues and under different stress conditions was analyzed. The results of the study are as follows:1. The gene structure and protein domain of PtPIF3a/ PtPIF3b, PtPIF4/ 5a/ PtPIF4/ 5b, PtPIF8a/ PtPIF8b and PtPIF9a/ PtPIF9b were found by the system evolution analysis, and the gene structure and the protein domain of PtPIFs were relatively conserved. And the predicted PtPIFs gene promoter region comprises a stress response element such as a hormone. It is shown that PtPIF can respond to changes of environmental factors, such as light.2. PtPIFs gene has tissue expression specificity and can respond to various abiotic stress. The expression of PtPIF1, PtPIF8a and PtPIF8b is mainly expressed in the mature leaves, and the expression of PtPIF1, PtPIF4/ 5a, PtPIF8b and PtPIF9b is down-regulated under the low temperature treatment, and the PtPIF10 is up-regulated; in the drought treatment, the expression amount of the PtPIF3b and the PtPIF4/ 5b is down-regulated, and the PtPIF8a is up-regulated; and under the salt treatment, the expression amount of the PtPIF4/ 5a is reduced, PtPIF3b and PtPIF9a are up-regulated. It is shown that PtPIFs can respond to different environmental stresses. To verify that PtPIFs can be combined with G-box, PtPIF3a is selected to be constructed on pGADT7-Rec2 vector, and the G-box sequence and the mutated G-box sequence are connected in series and cloned to the pHIS2 vector in triplicate. The results of single-hybridization experiments show that PtPIF3a can be combined with G-box and can not be combined with the mutated G-box. The binding capacity of Pt PIF3a indicates that PtPIFs has the potential to control the PIN through G-box. According to the results of the system evolution analysis, PtPIN1s were divided into two groups, one is PtPIN1a and PtPIN1b, and the other is PtPIN1c and PtPIN1d. The results of GUS staining show that the expression of PtPIN1a and PtPIN1b at the root tip of the adventitious root is stronger than that of PtPIN1c and PtPIN1d, and the expression difference is generated in the callus stage formed by the adventitious root. And is not affected by exogenous auxin. At low temperature, drought and salt stress, the expression of PtPIN1a/ Pt PIN1b and Pt PIN1c/ PtPIN1d is also similar. Under the low temperature treatment, the expression of PtPIN1b was down-regulated, and the expression of PtPIN1b and PtPIN1c was up-regulated under the drought treatment, and the expression of PtPIN1d was down-regulated under the treatment of salt. The difference of PtPIN1s in response to stress indicates that the PIN protein of the woody plant is diverse in evolution and function. The gene structure and conserved domain of PtPIF gene of poplar were systematically analyzed in this study, and the expression patterns of PtPIF members under different tissues and at low temperature, drought and NaCl stress were detected. At the same time, the different expression patterns of PtIN1s in the formation of adventitious roots and different abiotic stresses are demonstrated. The binding capacity of PtPIF3a with G-box indicates that PtPIFs may interact with the PIN. The results of the study provide a basis for the in-depth study of the mechanism of PtPIF and PtIN1s in the control of plant growth and development in different environments.
【学位授予单位】：中国林业科学研究院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;S792.11
【参考文献】
相关期刊论文 前1条
1 苏晓华,黄秦军,张冰玉,张香华;中国杨树良种选育成就及发展对策[J];世界林业研究;2004年01期
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