菊花磷脂酶Dα基因的耐逆表达特性
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摘要
本研究设计了切花菊‘神马’磷脂酶Dα基因(CmPLDα)的特异性引物,利用荧光定量的方法分析了CmPLDα的组织表达特性及其在非生物胁迫、脱落酸(ABA)、黑斑病接种处理下的表达特性,研究了菊花CmPLDα在胁迫应答中的作用。结果表明:CmPLDα在根、茎、叶、花等组织均有表达,在茎、叶、花中表达量较高,在根中表达较低;CmPLDα的表达几乎不受低温影响,整体上受机械损伤抑制,受高温明显抑制,受缺磷显著诱导,盐胁迫诱导了胁迫后期CmPLDα的增加,CmPLDα受ABA处理的快速诱导;黑斑病菌接种2 d后也可诱导CmPLDα的表达。由此可知,CmPLDα为组成型表达,该基因参与了菊花多种胁迫过程,包括温度、缺磷、ABA信号及黑斑病等。
In present study,the primers specific to CmPLDα from chrysanthemum ‘Jinba'was designed for qRT-PCR analysis of CmPLDα expression profiles in different tissues and in response to abiotic stresses,and exogenous abscisic acid(ABA) and leaf black spot( Alternaria alternata) inoculation,and the roles of CmPLDα in stress responses were disclosed. Result showed that CmPLDα expressed in root,stem,leaf and flower at higher levels in stem,leaf and flower than in the root. Expression of CmPLDα was little affected by cold while generally inhibited by mechanical wound. Heat stress significantly decreased the expression of CmPLDα. Pi deficiency significantly induced the expression of CmPLDα. CmPLDα expression was induced in the late period of salt treatment but induced quickly by ABA treatment. The expression of CmPLDα increased two days after leaf black spot inoculation. It was suggested that expression of CmPLDα was constitutive,and the gene was involved in a variety of physiological processes including temperature stress,Pi deficiency stress,ABA-induced signals and leaf black spot inoculation.
In present study,the primers specific to CmPLDα from chrysanthemum ‘Jinba'was designed for qRT-PCR analysis of CmPLDα expression profiles in different tissues and in response to abiotic stresses,and exogenous abscisic acid(ABA) and leaf black spot( Alternaria alternata) inoculation,and the roles of CmPLDα in stress responses were disclosed. Result showed that CmPLDα expressed in root,stem,leaf and flower at higher levels in stem,leaf and flower than in the root. Expression of CmPLDα was little affected by cold while generally inhibited by mechanical wound. Heat stress significantly decreased the expression of CmPLDα. Pi deficiency significantly induced the expression of CmPLDα. CmPLDα expression was induced in the late period of salt treatment but induced quickly by ABA treatment. The expression of CmPLDα increased two days after leaf black spot inoculation. It was suggested that expression of CmPLDα was constitutive,and the gene was involved in a variety of physiological processes including temperature stress,Pi deficiency stress,ABA-induced signals and leaf black spot inoculation.
引文
李燕,薛立,吴敏.2007.树木抗旱机理研究进展.生态学杂志,26(11):1857-1866.
徐呈祥.2012.提高植物抗寒性的机理研究进展.生态学报,32 (24):7966-7980.
杨鹍,傅亚萍,张玉满,等.2008.水稻OsPLD3和OsPLD4基因及其启动子.生物工程学报,24(3):368-375.
赵鹏,王道龙.2011.磷脂酶D对拟南芥抗冻性的影响.中国农学通报,27(6):107-111.
de Torres Zabela M,Fernandez-Delmond I,Niittyla T,et al.2002.Differential expression of genes encoding Arabidopsis phospholipases after challenge with virulent or avirulent Pseudomonas isolates.Molecular Plant-Microbe Interactions,15:808-816.
Fan L,Zheng SQ,Wang XM.1997.Antisense suppression of phospholipase Dαretards abscisic acid-and ethylene-promoted senescence of postharvest Arabidopsis leaves.The Plant Cell,9:2183-2196.
Gao HS,Song AP,Zhu XR,et al.2012.The heterologous expression in Arabidopsis of a chrysanthemum Cys2/His2 zinc finger protein gene confers salinity and drought tolerance.Planta,235:979-993.
Hong YY,Pan XQ,Welti R,et al.2008.Phospholipase Dα3is involved in the hyperosmotic response in Arabidopsis.The Plant Cell,20:803-816.
Li MY,Welti R,Wang XM.2006.Quantitative profiling of Arabidopsis polar glycerolipids in response to phosphorus starvation.Roles of phospholipases Dζ1 and Dζ2 in phosphatidylcholine hydrolysis and digalactosyldiacylglycerol accumulation in phosphorus-starved plants.Plant Physiology,142:750-761.
Liu QL,Xu KD,Ma N,et al.2010.Isolation and functional characterization of DgZFP:A gene encoding a Cys2/His2-type zinc finger protein in chrysanthemum.Molecular Biology Reports,37:1137-1142.
Livak KJ,Schmittgen TD.2001.Analysis of relative gene expression data using real-time quantitative PCR and the2-ΔΔCTMethod.Methods,25:402-408.
Mishkind M,Vermeer JEM,Darwish E,et al.2009.Heat stress activates phospholipase D and triggers PIP2accumulation at the plasma membrane and nucleus.The Plant Journal,60:10-21.
Mishra G,Zhang WH,Deng F,et al.2006.A bifurcating pathway directs abscisic acid effects on stomatal closure and opening in Arabidopsis.Science,312:264-266.
Moreno-Pérez AJ,Martínez-Force E,Garcés R,et al.2010.Phospholipase Dalpha from sunflower(Helianthus annuus):Cloning and functional characterization.Journal of Plant Physiology,167:503-511.
Nakazawa Y,Sato H,Uchino M,et al.2006.Purification,characterization and cloning of phospholipase D from peanut seeds.The Protein Journal,25:212-223.
Song A,Lu J,Jiang J,et al.2012.Isolation and characterisation of Chrysanthemum crassum SOS1,encoding a putative plasma membrane Na+/H+antiporter.Plant Biology,14:706-713.
Testerink C,Munnik T.2005.Phosphatidic acid:A multifunctional stress signaling lipid in plants.Trends in Plant Science,10:368-375.
Tong Z,Hong B,Yang YJ,et al.2009.Overexpression of two chrysanthemum DgDREB1 group genes causing delayed flowering or dwarfism in Arabidopsis.Plant Molecular Biology,71:115-129.
Uraji M,Katagiri T,Okuma E,et al.2012.Cooperative function of PLDδand PLDα1 in abscisic acid-induced stomatal closure in Arabidopsis.Plant Physiology,159:450-460.
Wan SB,Tian L,Tian RR,et al.2009.Involvement of phospholipase D in the low temperature acclimation-inducedthermotolerance in grape berry.Plant Physiology and Biochemistry,47:504-510.
Wang CX,Zien CA,Afitlhile M,et al.2000.Involvement of phospholipase D in wound-induced accumulation of jasmonic acid in Arabidopsis.The Plant Cell,12:2237-2246.
Wang XM,Xu LW,Zheng L.1994.Cloning and expression of phosphatidylcholine-hydrolyzing phospholipase D from Ricinus communis L.The Journal of Biological Chemistry,269 :20312-20317.
Wang XM.2005.Regulatory functions of phospholipase D and phosphatidic acid in plant growth,development,and stress responses.Plant Physiology,139:566-573.
Welti R,Li WQ,Li MY,et al.2002.Profiling membrane lipids in plant stress responses.The Journal of Biological Chemistry,277:31994-32002.
Whitaker BD,Smith DL,Green KC.2001.Cloning,character-ization and functional expression of a phospholipase Dalpha cDNA from tomato fruit.Physiologia Plantarum,112:87-94.
Zhang Q,Lin F,Mao TL,et al.2012.Phosphatidic acid regulates microtubule organization by interacting with MAP65-1in response to salt stress in Arabidopsis.The Plant Cell,24 :4555-4576.
Zhao J,Devaiah SP,Wang CX,et al.2013.Arabidopsis phospholipase Dβ1 modulates defense responses to bacterial and fungal pathogens.New Phytologist,199:228-240.
徐呈祥.2012.提高植物抗寒性的机理研究进展.生态学报,32 (24):7966-7980.
杨鹍,傅亚萍,张玉满,等.2008.水稻OsPLD3和OsPLD4基因及其启动子.生物工程学报,24(3):368-375.
赵鹏,王道龙.2011.磷脂酶D对拟南芥抗冻性的影响.中国农学通报,27(6):107-111.
de Torres Zabela M,Fernandez-Delmond I,Niittyla T,et al.2002.Differential expression of genes encoding Arabidopsis phospholipases after challenge with virulent or avirulent Pseudomonas isolates.Molecular Plant-Microbe Interactions,15:808-816.
Fan L,Zheng SQ,Wang XM.1997.Antisense suppression of phospholipase Dαretards abscisic acid-and ethylene-promoted senescence of postharvest Arabidopsis leaves.The Plant Cell,9:2183-2196.
Gao HS,Song AP,Zhu XR,et al.2012.The heterologous expression in Arabidopsis of a chrysanthemum Cys2/His2 zinc finger protein gene confers salinity and drought tolerance.Planta,235:979-993.
Hong YY,Pan XQ,Welti R,et al.2008.Phospholipase Dα3is involved in the hyperosmotic response in Arabidopsis.The Plant Cell,20:803-816.
Li MY,Welti R,Wang XM.2006.Quantitative profiling of Arabidopsis polar glycerolipids in response to phosphorus starvation.Roles of phospholipases Dζ1 and Dζ2 in phosphatidylcholine hydrolysis and digalactosyldiacylglycerol accumulation in phosphorus-starved plants.Plant Physiology,142:750-761.
Liu QL,Xu KD,Ma N,et al.2010.Isolation and functional characterization of DgZFP:A gene encoding a Cys2/His2-type zinc finger protein in chrysanthemum.Molecular Biology Reports,37:1137-1142.
Livak KJ,Schmittgen TD.2001.Analysis of relative gene expression data using real-time quantitative PCR and the2-ΔΔCTMethod.Methods,25:402-408.
Mishkind M,Vermeer JEM,Darwish E,et al.2009.Heat stress activates phospholipase D and triggers PIP2accumulation at the plasma membrane and nucleus.The Plant Journal,60:10-21.
Mishra G,Zhang WH,Deng F,et al.2006.A bifurcating pathway directs abscisic acid effects on stomatal closure and opening in Arabidopsis.Science,312:264-266.
Moreno-Pérez AJ,Martínez-Force E,Garcés R,et al.2010.Phospholipase Dalpha from sunflower(Helianthus annuus):Cloning and functional characterization.Journal of Plant Physiology,167:503-511.
Nakazawa Y,Sato H,Uchino M,et al.2006.Purification,characterization and cloning of phospholipase D from peanut seeds.The Protein Journal,25:212-223.
Song A,Lu J,Jiang J,et al.2012.Isolation and characterisation of Chrysanthemum crassum SOS1,encoding a putative plasma membrane Na+/H+antiporter.Plant Biology,14:706-713.
Testerink C,Munnik T.2005.Phosphatidic acid:A multifunctional stress signaling lipid in plants.Trends in Plant Science,10:368-375.
Tong Z,Hong B,Yang YJ,et al.2009.Overexpression of two chrysanthemum DgDREB1 group genes causing delayed flowering or dwarfism in Arabidopsis.Plant Molecular Biology,71:115-129.
Uraji M,Katagiri T,Okuma E,et al.2012.Cooperative function of PLDδand PLDα1 in abscisic acid-induced stomatal closure in Arabidopsis.Plant Physiology,159:450-460.
Wan SB,Tian L,Tian RR,et al.2009.Involvement of phospholipase D in the low temperature acclimation-inducedthermotolerance in grape berry.Plant Physiology and Biochemistry,47:504-510.
Wang CX,Zien CA,Afitlhile M,et al.2000.Involvement of phospholipase D in wound-induced accumulation of jasmonic acid in Arabidopsis.The Plant Cell,12:2237-2246.
Wang XM,Xu LW,Zheng L.1994.Cloning and expression of phosphatidylcholine-hydrolyzing phospholipase D from Ricinus communis L.The Journal of Biological Chemistry,269 :20312-20317.
Wang XM.2005.Regulatory functions of phospholipase D and phosphatidic acid in plant growth,development,and stress responses.Plant Physiology,139:566-573.
Welti R,Li WQ,Li MY,et al.2002.Profiling membrane lipids in plant stress responses.The Journal of Biological Chemistry,277:31994-32002.
Whitaker BD,Smith DL,Green KC.2001.Cloning,character-ization and functional expression of a phospholipase Dalpha cDNA from tomato fruit.Physiologia Plantarum,112:87-94.
Zhang Q,Lin F,Mao TL,et al.2012.Phosphatidic acid regulates microtubule organization by interacting with MAP65-1in response to salt stress in Arabidopsis.The Plant Cell,24 :4555-4576.
Zhao J,Devaiah SP,Wang CX,et al.2013.Arabidopsis phospholipase Dβ1 modulates defense responses to bacterial and fungal pathogens.New Phytologist,199:228-240.