摘要
1、本研究从小麦基因芯片结果中选取一个与Wali6相似在盐胁迫24 h表达显著提高的基因(Gene ID:TC248207),以山融3号小麦叶片实验材料,进行了cDNA克隆,并命名为TaUES(Upregulated Expression under Saline-stress in wheat)。进一步序列分析表明,TaUES编码蛋白含有97个氨基酸,其相对分子量为10.3 kD,等电点为7.52,不含保守结构域,含有信号肽,有可能是分泌蛋白。
基因枪转化洋葱表皮细胞的瞬时表达表明了TaUES-GFP融合蛋白的亚细胞定位无特异性,在细胞核与细胞质中均有表达,经过PI染色后,显示其定位于细胞核,关于TaUES的精确定位还需使用多种细胞器染料进行复染来进一步研究确定。
对TaUES的半定量RT-PCR的初步分析表明,根中TaUES基因经过NaCl处理24 h后被诱导表达,当小麦从含NaCl培养液中转移到正常培养液中恢复培养24 h后,其表达量逐渐降低,48 h后基本检测不到,根中TaUES基因属于晚期诱导表达类型。叶片中TaUES基因经过NaCl处理12 h后表达量最大,当小麦从含NaCl培养液中转移到正常培养液中后,其表达量能恢复到基准水平,叶片中TaUES基因属于晚期表达类型。
小麦根中TaUES基因经过PEG处理的表达模式呈现出与上述表达不同的模式,12 h后被诱导表达出现一个短暂的峰值,处理48 h后达到最大值;当小麦从含PEG培养液中转移到正常培养液中后,其表达量逐渐降低至检测不到,因此干旱胁迫根中TaUES基因属于晚期诱导表达类型。叶片中TaUES基因经过PEG处理后,表达量持续增大,在处理6 h后达到最大值,并在后续的处理中一直维持在这个较高的表达水平上;当小麦从含PEG培养液中转移到正常培养液中恢复培养后,其表达量逐渐降低直至检测不到,因此干旱胁迫叶片中TaUES基因属于早期诱导表达类型。
为了研究TaUES基因的功能,我们构建了植物过量表达表达载体,并进行了烟草和拟南芥转化。过量表达载体成功转化到烟草中,并得到T1代株系。拟南芥T0代正在筛选中。
2、小麦的生长发育、对外来信号的传导以及对逆境胁迫的应答反应构成了一个庞大而复杂的网络系统,其中丝氨酸/苏氨酸蛋白激酶的调控具有举足轻重的作用。从基因芯片结果中选取一个在盐胁迫24 h表达显著提高的基因(Gene ID:TC264050),以山融3号小麦叶片实验材料,克隆命名为TaSTK(Ta Ser/Thr Kinase)的cDNA,我们对其进行了序列分析和初步的功能验证。TaSTK编码蛋白含有453个氨基酸,其相对分子量为50.8 kD,等电点为767,第19到275氨基酸之间含有S_TKc保守结构域。预测该蛋白没有信号肽,可能定位于细胞质。
对TaSTK进行半定量RT-PCR的结果显示,盐胁迫和干旱胁迫影响TaSTK的表达,在小麦根和叶片中受到胁迫时均有表达并且表达量增加,我们认为TaSTK可能参与了对盐、干旱胁迫应答反应,并且属于早期表达类型
小麦根中TaSTK基因经过NaCl处理3 h后表达量开始增大,当小麦从含NaCl培养液中转移到正常培养液中48 h后,其表达量能恢复到基准水平;叶片中TaSTK基因经过NaCl处理后表现出与根中相似的表达模式,经过NaCl处理24 h后表达量达到最大。
小麦根中TaSTK基因经过PEG处理6 h后表达量开始增大,叶片中TaSTK基因经过PEG处理0.5 h后表达量开始增大,当小麦从含PEG培养液中转移到正常培养液中48 h后,根和叶片中其表达量能恢复到基准水平。
为了研究TaSTK的功能,构建了过量表达载体,并进行了烟草和拟南芥转化。过量表达载体通过农杆菌介导叶盘法成功转化到烟草中,并得到T1代株系。拟南芥T0代正在筛选中。同时也构建了TaSTK-GFP融合蛋白亚细胞定位载体,实验尚在进行中。
本实验从小麦中分离克隆出TaUES和TaSTK两个基因,并进行序列分析和功能验证,结果显示这两个基因可能参与盐和干旱胁迫反应。这为理解小麦响应干旱和盐胁迫等逆境胁迫的分子机理提供了实践基础,同时也为抗逆小麦的基因改良提供了理论依据。
1 We chose a gene similar to Wali6 (Gene ID: TC248207) whose expression level increased dramatically after 24 h of NaCl treatment from the results of gene chip, isolated a gene from the leaf of the somatic hybrid Shanrong No.3 by RT-PCR and named as TaUES (Upregulated Expression under Saline-stress in wheat). Sequence analysis shows that the TaUES cDNA has a single open reading frame (ORF) of 291 bp encoding a protein of 97 amino acids with an estimated molecular mass of 10.3 kD and an isoelectric point of 7.52.
The eatimated protein doesnot contain conserved domains, has a signal peptide and is possible to be a secreted protein. However, transient expression analysis in onion epidermal cells by genegun indicated TaUES-GFP fuse protein had no specific subcellular localization. Although we used PI to stain onion epidermal cells which had expressed TaUES-GFP fuse protein and the results showed it sulocated in the nucleus, we still needed to use a variety of dye staining to determine its precise location.
The expression patterns of TaUES in the leaf and root of wheat were analyzed by semiquantitative RT-PCR. It was found that the expression level of TaUES was high effected by salt and drought stress, and we concluded that TaUES was involved in responses to salt and drought stress. The expression of TaUES in root was induced slightly after 24 h of NaCl treatment, and when the roots were removed from medium containing NaCl, the expression of TaUES decreased to nearly zero. The expression of TaUES in leaf increased dramatically after 12 h of NaCl treatment, and when the roots were removed from medium containing NaCl, the expression of TaUES returned to pretreatment levels, showing that TaUES in leaf belonged to“late”expression type.
In contrast to above expression patterns, TaUES in the root which was treated by PEG had a complex pattern of induction, with a transient peak of expression after 12 h of PEG treatment and a second increase after 48 h. When the roots were removed from medium containing PEG, the expression of TaUES decreased to nearly zero. The expression level of TaUES in the leaf which was treated by PEG came to maximum and maintained at this higher level in the subsequent processing. When the roots were removed from medium containing PEG, the expression of TaUES returned to zero, showing that TaUES in leaf belonged to“early”expression type.
To further study the function of TaUES gene, we constructed a constitutive expression vector. Over-expression vector was transformed into tobacco and Arabidopsis. Now we obtained T1 line of tobacco and T0 line of Arabidopsis.
2 Wheat growth, development, signal conduction and stress responses form a complex and large network, in which serine/threonine protein kinase plays an important role in the regulation. We chose a gene (Gene ID: TC264050) whose expression level increased dramatically after 24 h of NaCl treatment from the results of gene chip. We isolated a gene from the leaf of the somatic hybrid Shanrong No.3 by RT-PCR and named as TaSTK. Sequence analysis shows that the TaSTK encodes a 50.8 kD protein with a calculated pI of 5.3 and has a S_TKc conserved domain from 19 to 279 amino acids. The eatimated protein has no signal peptide and sublocates in cytoplasm.
The expression patterns of TaSTK in the leaf and root of wheat were analyzed by semiquantitative RT-PCR. It was found that the expression level of TaSTK was high effected by salt and drought stress, and we concluded that TaSTK was involved in responses to salt and drought stress. The expression of TaSTK in root was induced slightly after 3 h of NaCl treatment, and when the roots were removed from medium containing NaCl, the expression of TaSTK decreased to nearly basal levels, which indicated that TaSTK in root fit the category of“eraly”induction. TaSTK mRNA in leaf followed the expression pattern of that in root, with maximal induction 24 h after NaCl exposure.
TaSTK in root and leaf showed simple induction, which the expression of TaSTK in root reached maximal level after 6h PEG exposure and that of TaSTK in root reached maximal level after 0.5 h PEG exposure, retumed to near basal levels when the roots were removed from medium containing PEG.
Overexpression vector was constructed to examine the function of TaSTK. By using Agrobacterium mediated leaf-disk tobacco transformation, the recombinant was transformed to Nicotiana tabacum L. Over-expression vector was also transformed into Arabidopsis. The transgenetic strains are screening.
In summary, we have isolated two wheat genes, TaUES and TaSTK, and analyzed their physiological functions. These results will provide practical evidence to understand the mechanism of wheat resistant to salt and drought stress and supply theoretical mechanism to improve wheat salt and drought resistance.
引文
[1] Kimberley C S, Richard C C. Five genes induced by aluminum in wheat (Triticum aestivum L.) roots [J]. Plant Physiol, 1993, 103: 855-861
[2] Keith D R, Kimberley C S, Richard C C. wali6 and wali7: genes induced by aluminum in wheat (Triticum aestivum L.) roots [J]. Plant Physiol, 1994, 105: 1455-1456
[3] Kawashima I, Inokuchi Y, Chino M, Kimura M, Shimizu N. Isolation of a gene for a metallothionein-like protein from soybean [J]. Plant Cell Physiol, 1991, 32: 913-916
[4] Evans I M, Gatehouse L N, Gatehouse J A, Robinson N J, Crciy R R D. A gene from pea (Pisum sativum L.) with homology to metallothionein genes [J]. FEBS Lett, 1990, 262: 29-32
[5] de Miranda J R, Thomas M A, Thurman D A, Tomsett A B. Metallothionein genes from the flowering plant Mimulus guttatus [J]. FEBS Lett, 1990, 260: 277-280
[6] de Framond A J. A metallothionein-like gene from maize (Zea mays): cloning and characterization [J]. FEBS Lett, 1991, 290: 103-106
[7] Okumura N, Nishizawa N-K, Umehara Y, Mori S. An iron deficiency-specific cDNA from barley roots having two homologous cysteine-rich MT domains [J]. Plant Mol Biol, 1991, 17: 531-533
[8] Berg J M. Potential metal-binding domains in nucleic acid binding proteins [J]. Science, 1986, 232: 485–487
[9] Laskowski M, Jr, Kato I. Protein inhibitors of proteinase [J]. Ann Rev Biochem, 1980, 49: 593-626
[10] Bode W, Huber R. Natural protein proteinase inhibitors and their interaction with proteinases [J]. Eur J Biochem, 1992, 204(2): 433-451
[11] Mello M O, Tanaka A S, Silva-Filho M C. Molecular evolution of Bowman-Birk type proteinase inhibitors in flowering plants [J]. Molecular Phylogenetics and Evolution, 2003, 27: 103-112
[12] Prakash B, Selvaraj S, Murthy M R, Sreerama Y N, Rao D R, Gowda L R. Analysis of the amino acid sequences of plant Bowman-Birk inhibitors [J]. J Mol Evol, 1996, 42(5): 560-569
[13] Li L, Zhao Y, Ma J L. Recent progress on key enzymes: PAL, C4H, 4CL of phenylalaninemetabolism pathway [J]. China Journal of Bioinformatics, 2007, 5(4): 187-189 (in Chinese with English Abstract)
[14] Alexandrov N N, Brover V V, Freidin S, Troukhan M E, Tatarinova T V, Zhang H, Swaller T J, Lu Y P, Bouck J, Flavell R B, Feldmann K A. Insights into corn genes derived from large-scale cDNA sequencing [J]. Plant Mol Biol, 2009, 69(1-2): 179-194
[15] Lu T, Piao X S. Advance in the effect of forsythia suspense extract on antioxidant activity [J]. Chinese Journal of Animal Science, 2009, 45(15): 57-60 (in Chinese with English Abstract)
[16] Tian G Z, Li H F, Qiu W F. Advances on research of plant peroxidases [J]. Journa l of Wuhan Botanica l Research, 2001, 19(4): 332-344 (in Chinese with English Abstract)
[17] Zhang K C, Jin Q, Cai Y P, Lin Y. Research progress of PAL and its control function of important secondav metabolltes [J]. Chinese agricultural science bulletin, 2008, 24(12): 59-62 (in Chinese with English Abstract)
[18] Tian X J, Qiu Z B, Liu X, Yue M. Effects of enhanced ultraviolet-B irradiance on the diurnal variation of flavonoids in wheat leaves [J]. Acta Scientiae Circumstantiae, 2007, 27(3), 516-521 (in Chinese with English Abstract)
[19] Chang T J, Zhu Z. Study advances of plant metallothionein-classification, characteristics and gene structure [J]. Biotechnology Bulletin, 2002, 3: 5-10 (in Chinese with English Abstract)
[20] Zhan Y, Yang C P. Study advance of metallothionein. Molecular Plant Breeding, 2006, 4(3): 73-78 (in Chinese with English Abstract)
[21] Xiao X X, Liu X H, Yang Z W, Wan Q, Zheng R, Wan Z J. Effect of aluminum stress on the content of protein and nucleic acid of longan (Dimocarpus longan) seedlings [J]. Silvae Sinicae, 2006, 42(10): 24-30 (in Chinese with English Abstract)
[22] Rakwal R, Kumar Agrawal G, Jwa N S. Characterization of a rice (Oryza sativa L.) Bowman-Birk proteinase inhibitor: tightly light regulated induction in response to cut, jasmonic acid, ethylene and protein phosphatase 2A inhibitors [J]. Gene, 2001, 263(1-2): 189-198
[23] Ezaki B, Gardner R C, Ezaki Y, Matsumoto H. Expression of aluminum-induced genes in transgenic arabidopsis plants can ameliorate aluminum stress and/or oxidative stress [J]. Plant Physiol, 2000, 122(3): 657-665
[24] Cao A Z, Li Q, Chen Y P, Zou X W, Wang X E, Chen P D. Screening resistance-related genes to powdery mildew in Haynaldia villosa using barley genechip and studying itsmechanism of resistance [J]. Acta Agronomic Sinica, 2006, 32(10): 1444-1452 (in Chinese with English Abstract)
[25] Hardie D G. Plant protein Serine/Threonine kinase: Classification and functions [J]. Annu. Rev. Plant Physiol. Plant Mol. Biol., 1999, 50: 97–131
[26] Bush D S. Calcium regulation in plant cells and its role in signaling [J]. Annu. Rev. Plant Physiol. PlantMol. Biol., 1995, 46: 95–122
[27] Harmon A C, Putnam-Evans, Cormier M J. A calcium-dependent but calmodulin-independent protein kinase from soybean [J]. Plant Physiol., 1987, 83: 830-837
[28] Halford N G, Hardie D G. SNF1-related protein kinases: global regulators of carbon metabolism in plants [J]. Plant Mol. Biol., 1998, 37: 735–748
[29] Hardie D G, Carling D, Carlson M. The AMP-activated/SNF1 protein kinase subfamily: metabolic sensors of the eu-karyotic cell [J]. Annu. Rev. Biochem., 1998, 67: 821–855
[30] Hrabak E M, Chan C W, Gribskov M, Harper J F, Choi J H, Sussman M R et al. The Arabidopsis CDPK-SnRK superfamily of protein kinase [J]. Plant Physiol., 2003, 132: 666-680
[31] Torii K U. Leucine-rich repeat receptor kinases in plants: structure, function, and signal transduction pathways [J]. Int. Rev. Cytol., 2004, 234: 1-46
[32] Walker J C, Zhang R. Relationship of a putative receptor protein kinase from maize to the S-locus glycoprotens of Brassica [J]. Nature, 1990, 345(6277): 743-746
[33] Navarro-Gochicoa M T, Camut S, Timmers A C, Niebel A, Herve C, Bono J J, Imberty A, Cullimore J V. Characterization of four lectin-like receptor kianses expressed in roots of Medicago truncatula, structure, location, regulation of expression, and potential role in the symbiosis with Sinorhizobium meliloti [J]. Plant Physiol., 2003, 133(4): 1893-1910
[34] Decreux A, Thomas A, Spies B, Brasseur R, Vancutsem P, Messiaen J. In vitro characterization of the homogalacturonan-binding domain of the wall-assosiated kinase WAKI using site-directed mutagenesis [J]. Phytochemistry, 2006, 67(11): 1068-1079
[35] Hu X, Reddy A S. Cloning and expression of a PR5-like protein from Arabidopsis: inhibition of fungal growth by bacterially expressed protein [J]. Plant Mol. Biol., 1997, 34(6): 949-959
[36] Jin P, Guo T, Becraft P W. The maize CR4 receptor-like kinase mediates a growth factor-like differentiation response [J]. Genesis, 2000, 27(3): 104-116
[37] Niu J S. Studies on plant and wheat protein kinases [J]. Acta Bot. Borea1.-Occident. Sin., 2003, 23 (1): 143-150
[38] Menges M, de Jager S M, Gruissem W, Murray J A H. Global analysis of the core cell cycle regulators of Arabidopsis identifies novel genes,reveals multiple and highly specific profiles of expression and provides a coherent model for plant cell cycle control [J]. Plant J, 2005, 41: 546-566
[39] Menges M, Murray J A H. Synchronous Arabidopsis suspension cultures for analysis of cell-cycle gene activity [J]. Plant J, 2002, 30:203-212
[40] Vandepoele K, Raes J, De Veylder L, Rouze P, Rombauts S, InzéD. Genome-wide analysis of core cell cycle genes in Arabidopsis [J]. Plant Cell, 2002, 14: 903-916
[41] Burssens S, Van Montagu M, InzéD. The cell cycle in Arabidopsis [J]. Plant Physiol Biochem, 1998, 36: 9-19
[42] Hashimoto J, Hirabayashi T, Hayano Y, Hata S, Ohashi Y, Suzuka I, Utsugi T, Toh-E A, Kikuchi Y. Isolation and characterization of cDNA clones encoding cdc2 homologuesfrom Oryza sati.a: a functional homologue and cognate variants [J]. Molecular and General Genetics, 1992, 233:10-16
[43] Setiady Y Y, Sekine M, Hariguchi N, Yamamoto T, Kouchi H, Shinmyo A. Tobacco mitotic cyclins: cloning, characterization, gene expression and functional assay [J]. Plant J, 1995, 8: 949-957
[44] Colasanti J, Tyers M, Sundaresan V. Identification and characterization of cDNA clones encoding a functional p34cdc2 homologue from Zea mays [J]. Proc Natl Acad Sci USA, 1991, 88:3377-3381
[45] Feiler H S, Jacobs T W. Cell division in higher plants: a cdc2 gene, its 34-kDa product, and histone H1 kinase activity in pea [J]. Proc Natl Acad Sci USA, 1990, 87:5397-5401
[46] Magyar Z, Meszaros T, Miskolczi P, Deak M, Feher A, Brown S, Kondorosi E, AthanasiadisA, Pongor S, Bilgin M. Cell cycle phase specificity of putative cyclin-dependent kinse variants in synchronized alfalfa cells [J]. Plant Cell, 1997, 9:223-235
[47] Graves P R, Roach P J. Role of COOH-terminal phosphorylation in the regulation of casein kinaseⅠδ[J]. Journal of Biological Chemistry, 1995, 270: 21689-21694
[48] Rychlik W Zagorski W. Purification and characterization of adenosine-3’,5’-phosphate-independent protein kinase from wheagerm [J]. Eur. J. Biochem., 1980, 106: 653-659
[49] Riera M, Peracchia G, de Nadal E, Arino J, Pages M. Maize protein kinase CK2: regulation and functionality of three beta regulatory subunits [J]. Plant J, 2001, 25(4): 365-374
[50] Litchfield D W. Protein kinase CK2: structure, regulation and role in cellular decisions of life and death [J]. Biochem J., 2003, 369(Pt 1): 1-15
[51] Kato K, Kidou S, Miura H. Molecular cloning and mapping of casein kinase 2 alpha and beta subunit genes in barley [J]. Genome, 2008, 51(3): 208-215
[52] Jonak C, Hirt H. Glycogen synthase kinase 3/SHAGGY-like kinases in plants: an emerging family with novel functions [J]. Trends Plant Sci, 2002, 7: 457?461
[53] Pay A, Jonak C, Bogre L, Meskiene I, Mairillger T, Szalay A, Heberle-Bors E, Hir H.The MsK family of alfalfa protein kinase genes encodes homologues of shaggy/glycogen synthase kinase-3 and shows differential expression patterns in plant, organs and development [J]. Plant J, 1993, 3: 847-856
[54] Bianchi M W, Guivarc’h D, Thomas M. Arabidopsis homologs of the shaggy and GSK-3 protein kinases: molecular cloning and functional expression in Escherichia coli [J]. Mol Gen Genet, 1994, 242: 337-345
[55] Chen G P, Ma W S, Huang Z J. Isolation and characterization of TaGSK1 involved in wheat salt tolerance [J]. Plant Science, 2003, 165(6): 1369-1375
[56] Kikuchi S, Satoh K, Nagata T. Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice [J]. Science, 2003, 301: 376-379
[57] Yang X G, Ma W S, Zhu Z G, Huang Z J, She Y Z. Plant GSKs: an emerging family with novel functions [J]. China BIiotechnology, 2006, 24(6): 25-27
[58] Urao T, Katagiri T, Mizoguchi T. Two genes that encode Ca2+-dependent protein kinase are induced by drought and high-salt sress in Arabodopsis thaliana [J]. Mol Gen Genet, 1994, 244: 331-340
[59] SaijoY, Hata S, KyozukaJ, Shimamoto K, Izui K. Over expression of a single Ca2+ dependent protein kinase confers both cold and salt/drought tolerance on rice plants [J]. Plant J, 2000, 23: 319-327
[60] Wang XQ, Wu W H. Involvement of a calcium-dependent protein kinase in ABA-regulationof stomatal movement [J]. Acta Bot Sinica, 1999, 41: 556-559
[61] Dunning F M, Sun W X, Jansen K L, Helft L, Bent A F. Identification and mutational analysis of Arabidopsis FLS2 leucine-rich repeat domain residues that contribute flagellin percepton [J]. Plant Cell, 2007, 19: 3297-3313
[62] Park C J, Peng Y, Chen X, Dardick C, Ruan D, Bart R, Canlas P E, Ronald P C. Rice XB15, a protein phosphatase 2c, negatively regulates cell death and Xa21-mediated innate immunity [J]. Plos. Biol., 2008, 6(9): 1910-1926
[63] Desikan R, Cheung M K, Bright J. ABA, hydrogenperoxide and nitric oxide signaling in stomatal guard cells [J]. J Exp Bot, 2004, 55(395): 205-212
[64] Liu Y, Zhang S. Phosphorylation of 1-aminocyclopropane-1-carboxylic acid synthase by MPK6, a stress responsive mitogen-activated protein kinase, induces ethylene biosynthesis in Arabidopsis [J]. Plant Cell, 2004, 16: 3386-3399
[65] Kim C Y, Liu Y, Thorne E T, Yang H, Fu Kushige H, Gassmann W, Hild Ebrand D, Sharpr E, Zhang S. Activation of a stress-responsive mitogen-activated protein kinase cascade induces the biosynthesis of ethylene in plants [J]. Plant Cell, 2003, 15: 2707-2718
[66] Kovtun Y, Chiu W, Tena G, Sheen J. Functional analysis of oxidative stress-activated mitogen-activated protein kinase cascade in plants [J]. Proc. Natl. Acad. Sci. USA, 2000, 97: 2940-2945
[67] Mockaitis K, Howell S H. Auxin induces mitogenic activated protein kinase (MAPK) activation in roots of Arabidopsis seedlings [J]. Plant J., 2000, 24: 785-796
[68] Hardtke C S, Deng X W. The cell biology of the COP/DET/FUS proteins. Regulating proteolysis in photomor phogenesis and beyond [J]? Plant Physiol., 2000, 124(4): 1548-1557
[69] Li J, Nam K H. Regulation of brassinosteroid signaling by a GSK/SHAGGY-like kinase [J]. Science, 2002, 295: 1299-1301
[70] Bogre L, Calderini O, Binarova P, Mattauch M, Till S, Kiegerl S, Jona K C, Pollaschek C, Barker P, Huskisson N S, Hirt H, Heberl E-Bors E. A MAP kinase is activated late in plant mitosis and becomes localized to t he plane of cell division [J]. Plant Cell, 1999, 11: 101-113
[71] Soyano T, Nishihama R, Morikiyo K, Ishikawa M, Machida Y. NQK1/NtMEK1 is a MAPKK that acts in the NPK1 MAPKKK mediated MAPK cascade and is required for plant cytokinesis [J]. Genes Dev., 2003, 17: 1055-1067
[72] Sorrell DA, Marchbank A, McMaho K, Dickinson J R, Rogers H J, Francis D. A WEE1 homologue from Arabidopsis thaliana [J].Planta, 2202, 215:518-522
[73] Romeis T, Piedras P, Jones J D G. Resistance gene-dependent activation of a cacium-dependent protein kinase in the plant defense response [J]. Plant Cell, 2000, 12: 803-816
[74] Lovas A, BimbóA, SzabóL, Bánfalvi Z. Antisense repression of StubGAL83 affects root and tuber development in potato [J]. Plant J., 2003, 33(1): 139-147
[75] Hao L, Wang H, Sunter G, Bisaro D M. GeminivirusAL2 and L2 proteins interact with and inactivate SNF1 kinase [J]. The Plant Cell, 2003, 15(4): 1034-1048
[76] Mao X, Zhang H, Tian S, Chang X, Jing R. TaSnRK2.4, an SNF1-type serine/threonine protein kinase of wheat (Triticum aestivum L.), confers enhanced multistress tolerance in Arabidopsis [J]. J. Exp. Bot., 2010, 61(3): 683-396
[77] Liu W, Xu ZH H, Luo D, Xue H W. OsCKI1, a rice casein kinaseⅠ, plays significant roles in auxin related root development and functions of plant hormones [J]. Plant J., 2003, 36: 189-202
[78] Wu L Z, Zhao B C, Qi Z G, Ge R C, Ma W S, Shen Y Z, Huang Z J. Localization and function analysis of wheat glycogen syntheses kinase (TaGSK1) [J]. Scientia Agricultura Sinica, 2006, 39(4): 842-847
[79] Anil V S, Harmon A C, Rao S K. Spatio-temporal accumulation and activity of calcium-dependent protein kinases durng embryogenesis, seed development, and germination in sandalwood [J]. Plant Physiol., 2000,122: 1035-1044
[80] Nishiyama R, Mizuno H, Okada S, Yamaguchi T, Takenaka M, Fukuzama H, Ohyama K. Two mRNA species encoding calcium-dependent protein kinase are differentially expressed in sexual organs of Maechatia polymopha through alternative splicing [J]. Plant Cell Physiol., 1999, 40: 205-212
[81] Halford N G, Hey S, Jhurreea D. Metabolic signaling and carbon partitioning: role of Snf1-related (SnRK1) protein kinase [J]. J. Exp. Bot., 2003, 54: 467-475
[82] Gong D, Zhang C, Chen X, Gong Z, Zhu J K. Constitutive activation and transgenic evaluation of the function of an Arabidopsis PKS protein kinase [J]. J. Biol. Chem., 2002, 277(44): 42088-42096
[83] Guo Y, Xiong L, Song C P, Gong D, Halfter U, Zhu J K.A calcium sensor and its interacting protein kinase areglobal regulators of abscisic acid signaling in Arabidopsis [J]. Dev. Cell, 2002, 3(2): 233-244
[84] Kim K N, Cheong Y H, Grant J J, Pandey G K, Luan S. CIPK3, a calcium sensor-associated protein kinase that regulates abscisic acid and cold signal transduction in Arabidopsis [J]. The Plant Cell, 2003, 15(2): 411-423
[85] Dai Y, Wang H, Li B, Huang J, Liu X, Zhou Y, Mou Z, Li J. Increased expression of map kinase kinase7 causes deficiency in polar auxin transport and leads to plant architectural abnormality in Arabidopsis [J]. Plant Cell, 2006, 18: 308-320
[86] Nieva C, Busk P K, Domínguez-Puigjaner E, Lumbreras V, Testillano P S, Risue?o M C, Pagès M. Isolation and functional characterisation of two new bZIP maize regulators of the ABA responsive gene rab28 [J]. Plant Mol. Biol., 2005, 58(6): 899-914
[87] Marcelo C D, Peter W, Von Rechlinghausen I. Characterization of three novel members of the Arabidopsis SHAGGY-related protein kinase (ASK) multigene family [J]. Plant Mol. Biol., 1999, 39: 137-147
[88] Xia G M, Xiang F N, Zhou A F, Wang H, Chen H M. Asymmetric somatic hybridization between wheat (Triticum aestivum L.) and Agropyron elongatum (Host) Nevishi[J]. Theor. Appl. Genet., 2003, 107: 299-305
[89] Chen S Y, Xia G M, Quan T Y, Xiang F N. Studies on the salt-tolerance of F3-F6 hybrid Lines originated from somatic hybridization between common wheat and Thinopyrum ponticum[J]. Plant Sci., 2004, 167, 773-779
[90]Liu S W, Zhao S Y, Chen F G., Xia G M. Generation of novel high quality HMW-GS genes in two introgression lines of Triticum aestivum/Agropyron elongatum[J]. BMC Evol. Biol., 2007, 7: 76-83
[91] Wang M C, Peng, Z Y, Li C L, Xia G M. Proteomic analysis on a high salt tolerance introgression strain of Triticum aestivum/Thinopyrum ponticum[J]. Proteomics 8, 2008,1470-1489