楊樹PtPIF和PtPIN1s基因功能的初步分析
發(fā)布時(shí)間：2019-05-17 10:22
【摘要】：光敏色素相互作用因子(Phytochrome Interacting Factor,PIF)是堿性螺旋-環(huán)-螺旋(basic Helix-Loop-Helix,bHLH)轉(zhuǎn)錄因子家族的一員,參與了植物光信號(hào)傳導(dǎo)。擬南芥基因組共編碼7個(gè)PIFs,在種子萌發(fā)、下胚軸伸長(zhǎng)、植物蔽蔭反應(yīng)和葉片衰老等過(guò)程具有重要功能,通過(guò)調(diào)控生長(zhǎng)素的合成與分布是其影響植物生長(zhǎng)發(fā)育的重要途徑。木本植物具有不同于草本植物的生長(zhǎng)發(fā)育過(guò)程,因此對(duì)光信號(hào)的應(yīng)答也存在差異。研究楊樹中PIF基因功能有助于揭示樹木光信號(hào)響應(yīng)機(jī)制。本研究分析了楊樹Pt PIF基因家族成員的系統(tǒng)進(jìn)化關(guān)系、基因結(jié)構(gòu)、保守結(jié)構(gòu)域及不同組織和不同脅迫條件下的表達(dá)模式。生長(zhǎng)素運(yùn)輸載體PIN是植物特有的,細(xì)胞膜極性定位的PIN蛋白可能受到PIF調(diào)控生長(zhǎng)素的運(yùn)輸改變生長(zhǎng)素濃度梯度,從而影響植物生長(zhǎng)發(fā)育。毛果楊共有15個(gè)Pt PIN基因,相對(duì)于擬南芥發(fā)生了顯著擴(kuò)張。為了探究PtPIN1s在表達(dá)及功能上的差異,通過(guò)GUS染色分析了PtPIN1s的組織表達(dá)差異,并分析了PtPIN1s在不同組織和不同脅迫條件下的表達(dá)模式。研究結(jié)果如下:1.篩選得到10個(gè)PtPIF基因家族成員,是擬南芥7個(gè)PIF基因家族成員的1.43倍;系統(tǒng)進(jìn)化分析發(fā)現(xiàn)了4個(gè)同源基因?qū)?PtPIF3a/PtPIF3b、PtPIF4/5a/PtPIF4/5b、PtPIF8a/PtPIF8b和PtPIF9a/PtPIF9b);PtPIFs的基因結(jié)構(gòu)和蛋白結(jié)構(gòu)域相對(duì)保守;且預(yù)測(cè)PtPIFs基因啟動(dòng)子區(qū)包含激素等脅迫響應(yīng)元件。表明PtPIF能夠響應(yīng)環(huán)境因子的變化,比如光。2.PtPIFs基因具有組織表達(dá)特異性并能響應(yīng)多種非生物脅迫。Pt PIF1、PtPIF8a和PtPIF8b主要在成熟葉中表達(dá),PtPIF3a和Pt PIF9b主要在幼葉中表達(dá);低溫處理下,PtPIF1、PtPIF4/5a、PtPIF8b和PtPIF9b的表達(dá)量下調(diào),PtPIF10上調(diào);干旱處理下,PtPIF3b和PtPIF4/5b的表達(dá)量下調(diào),PtPIF8a上調(diào);鹽處理下,PtPIF4/5a的表達(dá)量下調(diào),PtPIF3b和PtPIF9a上調(diào)。表明PtPIFs能夠響應(yīng)不同的環(huán)境脅迫。3.為了驗(yàn)證PtPIFs能夠與G-box結(jié)合,選取PtPIF3a構(gòu)建至pGADT7-Rec2載體上,將G-box序列與突變了的G-box序列以三次重復(fù)方式串聯(lián)并克隆至pHIS2載體上。酵母單雜交實(shí)驗(yàn)結(jié)果表明,PtPIF3a能夠與G-box結(jié)合,而不能與突變了的G-box結(jié)合。Pt PIF3a的結(jié)合能力表明PtPIFs有可能通過(guò)G-box調(diào)控PIN。4.根據(jù)系統(tǒng)進(jìn)化分析的結(jié)果將PtPIN1s分為兩類,一類是PtPIN1a和PtPIN1b,另一類是PtPIN1c和PtPIN1d。GUS染色結(jié)果發(fā)現(xiàn),PtPIN1a和PtPIN1b在不定根根尖處的表達(dá)強(qiáng)于PtPIN1c和PtPIN1d,這種表達(dá)差異產(chǎn)生于不定根形成的愈傷組織階段,且不受外源生長(zhǎng)素影響。在低溫、干旱及鹽脅迫下,PtPIN1a/Pt PIN1b和Pt PIN1c/PtPIN1d的表達(dá)有不同也有相似之處。低溫處理下,PtPIN1b的表達(dá)量下調(diào),PtPIN1c上調(diào);干旱處理下PtPIN1b和PtPIN1c的表達(dá)量上調(diào),PtPIN1d下調(diào);鹽處理下,PtPIN1d的表達(dá)量下調(diào)。PtPIN1s在響應(yīng)脅迫上的不同,表明木本植物PIN蛋白在進(jìn)化和功能上的多樣化。本研究預(yù)測(cè)了楊樹PtPIF基因家族成員,并對(duì)各成員的基因結(jié)構(gòu)、保守結(jié)構(gòu)域進(jìn)行了系統(tǒng)分析;檢測(cè)了PtPIF成員在不同組織及低溫、干旱和NaCl脅迫下的表達(dá)模式。同時(shí),證明了PtPIN1s在不定根形成及不同非生物脅迫下的不同表達(dá)模式。PtPIF3a與G-box的結(jié)合能力表明PtPIFs有可能與PIN發(fā)生相互作用。研究結(jié)果為深入探究楊樹PtPIF和PtPIN1s在不同環(huán)境下調(diào)控植物生長(zhǎng)發(fā)育的作用機(jī)制奠定了基礎(chǔ)。
[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.
【學(xué)位授予單位】：中國(guó)林業(yè)科學(xué)研究院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：Q943.2;S792.11
本文編號(hào)：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.
【學(xué)位授予單位】：中國(guó)林業(yè)科學(xué)研究院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：Q943.2;S792.11
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 蘇曉華,黃秦軍,張冰玉,張香華;中國(guó)楊樹良種選育成就及發(fā)展對(duì)策[J];世界林業(yè)研究;2004年01期
，本文編號(hào)：2479025
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