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平欧杂交榛CBF/DREB1转录因子ChaCBF1基因的克隆与功能分析

雷恒久 苏淑钗 马履一 马仲

雷恒久, 苏淑钗, 马履一, 马仲. 平欧杂交榛CBF/DREB1转录因子ChaCBF1基因的克隆与功能分析[J]. 北京林业大学学报, 2016, 38(10): 69-79. doi: 10.13332/j.1000-1522.20150528
引用本文: 雷恒久, 苏淑钗, 马履一, 马仲. 平欧杂交榛CBF/DREB1转录因子ChaCBF1基因的克隆与功能分析[J]. 北京林业大学学报, 2016, 38(10): 69-79. doi: 10.13332/j.1000-1522.20150528
LEI Heng-jiu, SU Shu-chai, MA Lü-yi, MA Zhong.. Cloning and functional analysis of ChaCBF1, a CBF/DREB1-like transcriptional factor from Corylus heterophylla × C. avellana.[J]. Journal of Beijing Forestry University, 2016, 38(10): 69-79. doi: 10.13332/j.1000-1522.20150528
Citation: LEI Heng-jiu, SU Shu-chai, MA Lü-yi, MA Zhong.. Cloning and functional analysis of ChaCBF1, a CBF/DREB1-like transcriptional factor from Corylus heterophylla × C. avellana.[J]. Journal of Beijing Forestry University, 2016, 38(10): 69-79. doi: 10.13332/j.1000-1522.20150528

平欧杂交榛CBF/DREB1转录因子ChaCBF1基因的克隆与功能分析

doi: 10.13332/j.1000-1522.20150528
基金项目: 

国家林业局重点项目“榛子良种选育与栽培关键技术研究”(2011-03)。

详细信息
    作者简介:

    雷恒久,博士。主要研究方向:经济林栽培与分子生物学。Email: leihengjiu123@163.com 地址:100083北京市海淀区清华东路35号北京林业大学林学院。
       责任作者: 苏淑钗,教授,博士生导师。主要研究方向:经济林栽培与育种。Email: sushuchai@sohu.com 地址:同上。

    雷恒久,博士。主要研究方向:经济林栽培与分子生物学。Email: leihengjiu123@163.com 地址:100083北京市海淀区清华东路35号北京林业大学林学院。
       责任作者: 苏淑钗,教授,博士生导师。主要研究方向:经济林栽培与育种。Email: sushuchai@sohu.com 地址:同上。

Cloning and functional analysis of ChaCBF1, a CBF/DREB1-like transcriptional factor from Corylus heterophylla × C. avellana.

  • 摘要: 为研究榛属植物的抗寒分子机理,以平欧杂交榛‘达维’为试材,采用同源克隆和RT-PCR技术克隆获得一个CBF/DREB1转录因子基因ChaCBF1,GenBank登录号为KT757373。该基因开放阅读框为666 bp,编码221个氨基酸,其相对分子质量为26.4 kDa,理论等电点为5.88。氨基酸序列比对结果显示ChaCBF1含有一个高度保守的AP2/ERF结构域,以及PKK/RPAGRxKFxETRHP和DSAWR等CBF蛋白特征序列。系统进化树分析发现ChaCBF1与垂枝桦BpCBF3蛋白的亲缘关系最近。通过基因枪轰击法使重组质粒p1302-ChaCBF1-GFP在洋葱表皮细胞瞬时表达,亚细胞定位结果显示ChaCBF1定位于洋葱表皮细胞的细胞核。通过实时荧光定量PCR检测了ChaCBF1基因在多种非生物胁迫下的表达模式,结果表明ChaCBF1基因的表达能够持续而强烈地响应低温信号,同时其表达也受干旱、高盐和ABA的诱导。通过农杆菌介导的花序侵染法将重组质粒p1301bar-ChaCBF1转化野生型拟南芥,对ChaCBF1基因的功能进行了研究。结果表明:过表达ChaCBF1的转基因拟南芥增强了对冷冻胁迫的耐受性,其成活率显著高于对照植株;转基因植株在正常和低温条件下能够显著上调冷响应基因RD29A和COR47的表达,并积累较高水平的游离脯氨酸和可溶性糖等渗透调节物质,从而提高其抗寒能力。研究结果表明,转录因子ChaCBF1基因在杂交榛冷响应途径中发挥重要作用。

     

  • [1] ZHENG W J. Records of Chinese trees[M]. Beijing:China Forestry Publishing House, 1998.
    [1] 郑万钧.中国树木志[M]. 北京:中国林业出版社, 1998.
    [2] CHEN X,WANG G X,LIANG L S, et al. Cloning and temporal-spatial expression of a CBF homolog associated with cold acclimation from Corylus heterophylla[J]. Scientia Silvae Sinicae,2012,48(1):167-172.
    [2] 陈新,王贵禧,梁丽松,等.平榛冷适应相关基因CBF的克隆及时空表达特性分析[J]. 林业科学,2012, 48(1):167-172.
    [3] 李凤光,娄汉平,高丹,等.平欧杂交榛引种初报[J]. 北方园艺,2009, 33(4):91-92.
    [3] LI F G, LOU H P, GAO D, et al. Preliminary study on introducing of hybrid hazel[J]. Northern Horticulture, 2009, 33(4):91-92.
    [4] 李春牛,董凤祥,王贵禧,等.平欧杂交榛抗抽条能力及抽条临界含水量研究[J]. 林业科学研究,2010,23(3):330-335.
    [4] LI C N,DONG F X,WANG G X, et al. Study on the tolerance and critical water capacity of shoot shriveling in hybrid hazelnuts[J]. Forest Research,2010,23(3):330-335.
    [5] 赵爽,苏淑钗,张兵,等.河北省平泉县平欧杂交榛越冬性研究[J]. 中南林业科技大学学报,2015,35(4):33-39.
    [5] ZHAO S,SU S C,ZHANG B, et al. Study on over-wintering survival rate of Corylus heterophylla × C.avellana in Pingquan, Hebei Province[J]. Journal of Central South University of Forestry Technology, 2015,35(4):33-39.
    [6] 陈新,王贵禧,徐丽,等.榛子甜菜碱醛脱氢酶基因BADH的克隆及在越冬过程中的表达特性分析[J]. 山东农业科学,2013,45(5):6-12.
    [6] CHEN X,WANG G X,XU L,et al. Cloning and expression analysis of betaine aldehyde dehydrogenase BADH gene from hzaelnut(Corylus heterophylla Fisch.) during overwintering period[J]. Shandong Agricultural Sciences,2013,45(5):6-12.
    [7] QIN F,SHINOZAKI K,YAMAGUCHI-SHINOZAKI K. Achievements and challenges in understanding plant abiotic stress responses and tolerance[J]. Plant Cell Physiology,2011,52(9):1569-1582.
    [7] SONG C N,QIAN J L,FANG J G,et al. Cloning, subcellular localization and expression analysis of SPL9 and SPL13 genes from Poncirus trifoliata[J]. Scientia Agricultura Sinica,2010,43(10):2105-2114.
    [8] ZHAO S J,SHI G A,DONG X C. Techniques of plant physiological experiment[M]. Beijing:Agricultural Scientific and Technical Press of China, 2002.
    [8] STOCKINGER E J,GILMOUR S J,THOMASHOW M F. Arabidopsis thaliana CBF1 encodes an AP2 domain-containing transcriptional activator that binds to the C-repeat/DRE, a cis-acting DNA regulatory element that stimulates transcription in response to low temperature and water deficit[J]. Proceeding of the National Academy of Sciences of the United States of America,1997,94(3):1035-1040.
    [9] PAN X W,LI Y C,LI X X. Differential regulatory mechanism of CBF regulon between Nipponbare(japonica) and 93-11(Indica) during cold acclimation[J]. Chinese Journal of Rice Science,2012,26(5):521-528.
    [9] LIU Q,KASUGA M,SAKUMA Y,et al. Two transcription factors, DREB1 and DREB2, with an EREBP/AP2 DNA binding domain separate two cellular signal transduction pathways in drought-and low-temperature-responsive gene expression, respectively, in Arabidopsis[J]. The Plant Cell,1998,10(8):1391-1406.
    [10] WEI J Y,ZHAO J,ZHAO S Q. Activation and regulation on the cold response pathway of ICE1-CBF in plants[J]. Biotechnology Bulletin,2015,31(6):8-12.
    [10] SAKUMA Y,LIU Q,DUBOUZET J G, et al. DNA-binding specificity of the ERF/AP2 domain of Arabidopsis DREBs, transcription factors involved in dehydration-and cold-inducible gene expression[J]. Biochemical Biophysical Research Communications,2002, 290(3):998-1009.
    [11] DUBOUZET J G,SAKUMA Y,ITO Y, et al. OsDREB genes in rice, Oryza sativa L., encode transcription activators that function in drought-, high-salt-and cold-responsive gene expression[J]. The Plant Journal,2003,33(4):751-763.
    [12] QIN F,SAKUMA Y,LI J, et al. Cloning and functional analysis of a novel DREB1/CBF transcription factor involved in cold-responsive gene expression in Zea mays L.[J]. Plant Cell Physiology, 2004,45(8):1042-1052.
    [13] YANG W,LIU X D,CHI X J,et al. Dwarf apple MbDREB1 enhances plant tolerance to low temperature, drought, and salt stress via both ABA-dependent and ABA-independent pathways[J]. Planta,2011,233(2):219-229.
    [14] KITASHIBA H,ISHIZAKA T,ISUZUGAWA K, et al. Expression of a sweet cherry DREB1/CBF ortholog in Arabidopsis confers salt and freezing tolerance[J]. Journal of Plant Physiology,2004,161(10):1171-1176.
    [15] NAVARRO M,AYAX C,MARTINEZ Y, et al. Two EguCBF1 genes overexpressed in Eucalyptus display a different impact on stress tolerance and plant development[J]. Plant Biotechnology Journal, 2011,9(1):50-63.
    [16] WELLING A,PALVA E T. Involvement of CBF transcription factors in winter hardiness in birch[J]. Plant Physiology, 2008,147(3):1199-1211.
    [17] WANG Z L,LIU J,GUO H Y, et al. Characterization of two highly similar CBF/DREB1-like genes, PhCBF4a and PhCBF4b, in Populus hopeiensis[J]. Plant Physiology and Biochemistry,2014,83:107-116.
    [18] BENEDICT C,SKINNER J S,MENG R, et al. The CBF1-dependent low temperature signaling pathway, regulon and increase in freeze tolerance are conserved in Populus spp.[J]. Plant, Cell Environment, 2006,29(7):1259-1272.
    [19] CHENG H,CAI H B,FU H T, et al. Functional characterization of Hevea brasiliensis CRT/DRE binding factor1 gene revealed regulation potential in the CBF pathway of tropical perennial tree[J/OL]. PLoS One,2015,10(9):e0137634. [2015-12-12]. http:∥dx.doi.org/ 10.1371/journal.pone.0137634. DOI: 10.1371/journal.pone.0137634.
    [20] MORRAN S,EINI O,PYVOVARENKO T, et al. Improvement of stress tolerance of wheat and barley by modulation of expression of DREB/CBF factors[J]. Plant Biotechnology Journal,2011,9(2):230-249.
    [21] CHANG S,PURYEAR J,CAIRNEY J. A simple and efficient method for isolating RNA from pine trees[J]. Plant Molecular Biology Reporter,1993,11(2):113-116.
    [22] 宋长年,钱剑林,房经贵,等. 枳SPL9和SPL13全长cDNA克隆、亚细胞定位和表达分析[J]. 中国农业科学,2010,43(10):2105-2114.
    [23] LIVAK K J,SCHMITTGEN T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCt method[J]. Methods,2001, 25(4):402-408.
    [24] CLOUGH S J,BENT A F. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana[J]. The Plant Journal,1998,16(6):735-743.
    [25] BATES L S,WALDREN R P,TEARE I D. Rapid determination of free proline for water-stress studies[J]. Plant and Soil,1973,39(1):205-207.
    [26] 赵世杰,史国安,董新纯.植物生理学实验指导[M]. 北京:中国农业科学技术出版社, 2002.
    [27] JAGLO K R,KLEFF S,AMUNDSEN K L, et al. Components of the Arabidopsis C-repeat/dehydration-responsive element binding factor cold-response pathway are conserved in Brassica napus and other plant species[J]. Plant Physiology,2001,127(3):910-917.
    [28] XIONG Y W,FEI S Z. Functional and phylogenetic analysis of a DREB/CBF-like gene in perennial ryegrass(Lolium perenne L.)[J]. Planta,2006,224(4):878-888.
    [29] LI D F,ZHANG Y Q,HU X N,et al. Transcriptional profiling of Medicago truncatula under salt stress identified a novel CBF transcription factor MtCBF4 that plays an important role in abiotic stress responses[J]. BMC Plant Biology,2011,11:109.
    [30] CONG L,CHAI T Y,ZHANG Y X. Characterization of the novel gene BjDREB1B encoding a DRE-binding transcription factor from Brassica juncea L.[J]. Biochemistry Biophysical Research Communications,2008,371(4):702-706.
    [31] TONG Z,HONG B,YANG Y J, et al. Overexpression of two chrysanthemum DgDREB1 group genes causing delayed flowering or dwarfism in Arabidopsis[J]. Plant Molecular Biology,2009,71(1):115-129.
    [32] OAKENFULL R J,BAXTER R,KINGHT M R. A C-repeat binding factor transcriptional activator(CBF/DREB1) from European bilberry(Vaccinium myrtillus) induces freezing tolerance when expressed in Arabidopsis thaliana[J/OL]. PLoS One, 2013,8(1):e54119. [2015-12-13]. http:∥dx.doi.org/ 10.1371/journal.pone.0054119. DOI: 10.1371/journal.pone.0054119.
    [33] VOGEL J T,ZARKA D G,VAN BUSKIRK H A, et al. Roles of the CBF2 and ZAT12 transcription factors in configuring the low temperature transcriptome of Arabidopsis[J]. The Plant Journal,2005,41(2):195-211.
    [34] ISHITANI M,MAJUMDER A L,BORNHOUSER A ,et al. Coordinate transcriptional induction of myo-inositol metabolism during environmental stress[J]. The Plant Journal,1996,9(4):537-548.
    [35] WANNER L A,JUNTTILA O. Cold-induced freezing tolerance in Arabidopsis[J]. Plant Physiology,1999,120(2):391-399.
    [36] TAJI T,OHSUMI C,IUCHI S, et al. Important roles of drought-and cold-inducible genes for galactinol synthase in stress tolerance in Arabidopsis thaliana[J]. The Plant Journal,2002,29(4):417-426.
    [37] NOMURA M,MURAMOTO Y,YASUDA S, et al. The accumulation of glycinebetaine during cold acclimation in early and late cultivars of barley[J]. Euphytica,1995,83(3):247-250.
    [38] ITO Y,KATSURA K,MARUYAMA K, et al. Functional analysis of rice DREB1/CBF-type transcription factors involved in cold-responsive gene expression in transgenic rice[J]. Plant Cell Physiology,2006,47(1):141-153.
    [39] 潘孝武,黎用朝,李小湘.CBF调节子在水稻品种日本晴和93-11低温驯化过程中的差异调控机制[J]. 中国水稻科学,2012,26(5):521-528.
    [40] ZHANG X,FOWLER S G,CHENG H, et al. Freezing-sensitive tomato has a functional CBF cold response pathway, but a CBF regulon that differs from that of freezing-tolerant Arabidopsis[J]. The Plant Journal,2004,39(6):905-919.
    [41] 魏俊燕,赵佳,赵仕琪. 植物ICE1-CBF冷反应通路的激活与调控研究进展[J]. 生物技术通报, 2015,31(6):8-12.
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  • 收稿日期:  2016-01-08
  • 刊出日期:  2016-10-29

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