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作物学报 ›› 2010, Vol. 36 ›› Issue (3): 401-409.doi: 10.3724/SP.J.1006.2010.00401

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

利用cDNA-AFLP技术分析小麦成株抗条锈性差异基因表达特征

张岗1,**，董艳玲1,**，夏宁1，张毅1，王晓杰1，屈志鹏1，李依民1，黄丽丽1，康振生1,2,*   

  1. 1西北农林科技大学植物保护学院, 陕西杨凌 712100; 2西北农林科技大学 / 陕西省农业分子生物学重点实验室, 陕西杨凌 712100
  • 收稿日期:2009-10-16 修回日期:2009-12-08 出版日期:2010-03-12 网络出版日期:2009-12-21
  • 通讯作者: 康振生, E-mail: kangzs@nwsuaf.edu.cn
  • 基金资助:

    本研究由国家自然科学基金重点项目（30930064），现代农业产业技术体系技术专项资金，高等学校学科创新引智计划项目（B07049）和国家“十一五”科技支撑计划项目（2006BAD08A05）资助。

cDNA-AFLP Analysis Reveals Differential Gene Expression in Wheat Adult-Plant Resistance to Stripe Rust

ZHANG Gang1,**,DONG Yan-Ling1,**,XIA Ning1,ZHANG Yi1,WANG Xiao-Jie1,QU Zhi-Peng1,LI Yi-Min1,HUANG Li-Li1,KANG Zhen-Sheng1,2,*   

  1. 1 College of Plant Protection, Northwest A&F University, Yangling 712100, China; 2 Shaanxi Provincial Key Laboratory of Molecular Biology for Agriculture / Northwest A&F University, Yangling 712100, China
  • Received:2009-10-16 Revised:2009-12-08 Published:2010-03-12 Published online:2009-12-21
  • Contact: KANG Zhen-Sheng, E-mail: kangzs@nwsuaf.edu.cn

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被引次数

31

摘要：

采用cDNA-AFLP技术，对成株抗条锈小麦品种兴资9104在成株期受条锈菌生理小种CY32侵染后5 d内9个时间点的基因表达谱进行了分析。共筛选64对引物，产生32 320个转录本(TDF)；用37对引物检测到2 201个(6.81%)差异TDF，其中926个TDF诱导表达，1 275个下调表达。经大规模克隆、测序分析，最终获得330个差异TDF，聚类分析得到259个EST (unigenes)，命名为aTaPST1至aTaPST259 (GenBank注册号：FL645754~FL646011和FL646262)。经BLASTX比对和功能分类分析，其中96条EST(37.07%)未找到同源性匹配，68条(26.25%)与未知功能蛋白同源性较高；其余95条ESTs主要涉及能量(11.20%)、基础代谢(4.63%)、转录调控(3.86%)、抗病与防御(3.86%)、蛋白质运输和储存(3.09%)、蛋白质合成和细胞生长(各2.32%)、以及信号转导(1.54%)等。选取抗病与防御、转录调控及信号转导类等相关的6个差异基因，qRT-PCR分析结果显示其表达模式符合cDNA-AFLP表达谱。小麦成株抗条锈性分子机制涉及植物多方面生理生化反应，包括抗病与防御、转录调控、蛋白质代谢、信号转导、以及非生物胁迫等多种途径相关基因的协同控制。

关键词: 小麦, 条锈菌, 抗病性, 基因表达, cDNA-AFLP, qRT-PCR

Abstract:

Wheat (Triticum aestivum L.) stripe rust is one of the most devastating diseases of wheat throughout the world. Adult plant resistance (APR) to stripe rust in wheat conferring durable resistance, thus, plays a pivotal role in the control of the disease. In the present study, to elucidate molecular mechanism of wheat APR to stripe rust, we conducted extensive transcription profiling of adult-plant wheat cultivar Xingzi 9104 infected by Puccinia striiformis Westend f. sp. tritici Erikss. pathotype CY32 using cDNA-AFLP technique. We analyzed transcription profiling of the incompatible reaction across nine sampling time points within five days after inoculation. Of the total 32,320 transcript derived fragments (TDFs) obtained using cDNA-AFLP with 64 primer pairs, 2201 (6.81%) displayed altered expression patterns after inoculation, of which 926 showed up-regulated and 1275 down-regulated. Three hundred and thirty differentially expressed TDFs produced reliable sequences after cloning and sequencing, of which 259 expressed sequence tags (ESTs) of unigenes were obtained after assembling, designated from aTaPST1 to aTaPST259, deposited in GenBank with accessions numbers from FL645754 to FL646011 and FL646262. BLASTX analyses and functional annotations were then performed and the results revealed that the 95 ESTs had predicted gene products mainly implicated in energy (11.20%), metabolism (4.63%), transcription (3.86%), disease/defense (3.86%), protein destination and storage (3.09%), protein synthesis and cell growth (each accounted for 2.32%), and signal transduction (1.54% of the sequenced total 259 ESTs). Six differential genes related to disease/defense, transcription, and signal transduction were chosen for further qRT-PCR expression patterns, which confirmed the cDNA-AFLP profiles. Our results indicated that wheat APR to stripe rust involved in multifaceted biochemical and physiological reactions, including concerted regulation of the genes involved in different pathways like disease/defense, transcription, protein metabolism, signal transduction, as well as abiotic stresses. These results provide information for further elucidation of molecular mechanism of wheat APR to stripe rust.

Key words: Wheat, Stripe rust fungus, Adult-plant resistance, Gene expression, cDNA-AFLP, qRT-PCR


[1] Chen X M. Epidemiology and control of stripe rust (Puccinia striiformis f. sp. tritici) on wheat. Can J Plant Pathol, 2005, 27: 314-337

[2] Bariana H S, McIntos A H. Genetics of adult plant stripe rust resistance in four Australian wheats and the French cultivar ‘Hyhride-de-Bersee’. Plant Breed, 1995, 114: 485-491

[3] Fu D L, Uauy C, Distelfeld A, Blechl A, Epstein L, Chen X M, Sela H, Fahima T, Dubcovsky J. A Kinase-START gene confers temperature-dependent resistance to wheat stripe rust. Science, 2009, 323: 1357-1360

[4] Moldenhauer J, Moerschbacher B M, van der Westhuizen A J. Histological investigation of stripe rust (Puccinia striiformis f. sp. tritici) development in resistant and susceptible wheat cultivars. Plant Pathol, 2006, 55: 469-467

[5] Moldenhauer J, Pretorius Z A, Moerschbacher B M, Prins R, van der Westhuizen A J. Histopathology and PR-protein markers provide insight into adult plant resistance to stripe rust of wheat. Mol Plant Pathol, 2008, 9: 137-145

[6] Melichar J P E, Berry S, Newell C, MacCormack R, Boyd L A. QTL identification and microphenotype characterization of the developmentally regulated yellow rust resistance in the UK wheat cultivar Guardian. Theor Appl Genet, 2008, 117: 391-399

[7] Mallard S, Negre S, Pouya S, Gaudeet D, Lu Z X, Dedryver R F. Adult plant resistance-related gene expression in ‘Camp Remy’ wheat inoculated with Puccinia striiformis. Mol Plant Pathol, 2008, 9: 213-225

[8] Coram T E, Settles M, Chen X M. Transcriptome analysis of high-temperature adult-plant resistance conditioned by Yr39 during the wheat-Puccinia striiformis f. sp. tritici interaction. Mol Plant Pathol, 2008, 9: 479-493

[9] Hulbert S H, Bai J, Fellers J P, Pacheco M G, Bowden R L. Gene expression patterns in near isogenic lines for wheat rust resistance gene Lr34/Yr18. Phytophathology, 2007, 97: 1083-1093

[10]Liu H-M(刘红梅), Liu T-G(刘太国), Xu S-C(徐世昌), Liu D-Q(刘大群), Chen W-Q(陈万权). Inheritance of yellow rust resistance in an elite wheat germplasm Xingzi 9104. Acta Agron Sin (作物学报), 2006, 32(11): 1742-1745 (in Chinese with English abstract)

[11]Bachem C W B, Hoeven R S, van der Bruijn S M. de Vreugdenhil D, Zabeau M, Visser R G F. Visualization of differential gene expression using a novel method of RNA fingerprinting based on AFLP analysis of gene expression during potato tuber development. Plant J, 1996, 9: 745-753

[12]Wang X J, Tang C L, Zhang G, Li Y C, Wang C F, Liu B, Qu Z P, Zhao J, Han Q M, Huang L L, Chen X M, Kang Z S. cDNA-AFLP analysis reveals differential gene expression in compatible interaction of wheat challenged with Puccinia striiformis f. sp. tritici. BMC Genomics, 2009, 10: 289

[13]Zadoks J C, Chang T T, Konzak C F. A decimal code for the growth stages of cereals. Weed Res, 1974, 14: 415-421

[14]Kang Z-S(康振生), Li Z-Q(李振岐).Discovery of a normal T. type new pathogenic strain to Lovrin10. J Northwest Agric Coll (西北农学院学报), 1984, 12(4): 18-28 (in Chinese with English abstract)

[15]Bevans M, Bancroft I, Bent E, Love K, Goodman H, Dean C, Bergkamp R, Dirkse W, Van Staveren M, Stiekema W, Drost L, Ridley P, Hudson S A, Patel K, Murphy G, Piffanelli P, Wedler H, Wedler E, Wambutt R, Weitzenegger T, Pohl T M, Terryn N, Gielen J, Villarroel R, De Clerck R, Van Montagu M, Lecharny A, Auborg S, Gy I, Kreis M, Lao N, Kavanagh T, Hempel S, Kotter P, Entian K-D, Rieger M, Schaeffer M, Funk B, Mueller-Auer S, Silvey M, James R, Montfort A, Pons A, Puigdomenech P, Douka A, Voukelatou E, Milioni D, Hatzopoulos P, Piravandi E, Obermaier B, Hilbert H, Düsterhöft A, Moores T, Jones J D G, Eneva T, Palme K, Benes V, Rechman S, Ansorge W, Cooke R, Berger C, Delseny M, Voet M, Volckaert G, Mewes H W, Klosterman S, Schueller C, Chalwatzis N. Analysis of 1.9 Mb of contiguous sequence from chromosome 4 of Arabidopsis thaliana. Nature, 1998, 391: 485-488

[16]Pfaffl M W. A new mathematical model for relative quantification in real-time RT-PCR. Nucl Acids Res, 2001, 29: e45

[17]Tao Y, Xie Z, Chen W, Glazebrook J, Chang H S, Han B, Zhu T, Zou G Z, Katagiri F. Quantitative nature of Arabidopsis responses during compatible and incompatible interactions with bacterial pathogen Pseudomonas syringae. Plant Cell, 2003, 15: 317-330

[18]Martin G B, Brommonschenkel S H, Chunwongse J, Frary A, Ganal M W, Spivey R, Wu T, Earle E D, Tanksley S D. Map-based cloning of a protein kinase gene conferring disease resistance in tomato. Science, 1993, 262: 1432-1436

[19]Brueggeman R, Rostoks N, Kudrna D, Kilian A, Han F, Chen J, Druka A, Steffenson B, Kleinhofs A. The barley stem rust-resistance gene Rpg1 is a novel disease-resistance gene with homology to receptor kinases. Proc Natl Acad Sci USA, 2002, 99: 9328-9333

[20]Butt A, Mousley C, Morris K, Beynon J, Can C, Holub E, Greenberg J T, Buchanan Wollaston V. Differential expression of a senescence-enhanced metallothionein gene in Arabidopsis in response to isolates of Peronospora parasitica and Pseudomonas syringae, Plant J, 1998, 16: 209-221

[21]Akashi K, Nishimura N, Ishida Y, Yokota A. Potent hydroxyl radical-scavenging activity of drought-induced type-2 metallothionein in wild watermelon. Biochem Biophys Res Commun, 2004, 323: 72-78

[22]Sharma Y K, Davis K R. The effects of ozone on antioxidant responses in plants. Free Radical Biol Med, 1997, 23: 480-488

[23]Craig K L, Tyers M. The F-box: A new motif for ubiquitin dependent proteolysis in cell cycle regulation and signal transduction. Prog Biophys Mol Biol, 1999, 72: 299-328

[24]Woo H R, Chung K M, Park J H, Oh S A, Ahn T, Hong S H, Jang S K, Nam H G. ORE9, an F-box protein that regulates leaf senescence in Arabidopsis. Plant Cell, 2001, 13: 1779-1790

[25]Kim H S, Delaney T P. Arabidopsis SON1 is an F-box protein that regulates a novel induced defense response independent of both salicylic acid and systemic acquired resistance. Plant Cell, 2002, 14: 1469-1482

[26]Devoto A, Nieto-Rostro M, Xie D, Ellis C, Harmston R, Patrick E. COI1 links jasmonate signalling and fertility to the SCF ubiquitin-ligase complex in Arabidopsis. Plant J, 2002, 32: 457-466

[27]Van den Burg H A, Tsitsigiannis D I, Rowland O, Lo J, Rallapalli G, MacLean D, Takken F L W, Jones J D G. The F-box protein ACRE189/ACIF1 regulates cell death and defense responses activated during pathogen recognition in tobacco and tomato. Plant Cell, 2008, 20: 697-719

[28]Van Nocker S, Ludwig P. The WD-repeat protein superfamily in Arabidopsis: conservation and divergence in structure and function. BMC Genomics, 2003, 4: 50

[29]Walker A R, Davison P A, Bolognesi-Winfield A C, James Celia M, Srinivasan N, Blundell T L, Esch J J, Marks M D, Gray J C. The TRANSPARENT TESTA GLABRA1 locus, which regulates trichome differentiation and anthocyanin biosynthesis in Arabidopsis, encodes a WD40 repeat protein. Plant Cell, 1999, 11: 1337-1350

[30]Huang J, Wang M M, Bao Y M, Sun S J, Pan L J, Zhang H S. SRWD: a novel WD40 protein subfamily regulated by salt stress in rice (Oryza sativa L.). Gene, 2008, 424: 71-79

[31]Jensen M K, Rung J H, Gregersen P L, Gjetting T, Fuglsang A T, Hansen M, Joehnk N, Lyngkjaer M F, Collinge D B. The HvNAC6 transcription factor: A positive regulator of penetration resistance in barley and Arabidopsis. Plant Mol Biol, 2007, 65: 137-150

[32]Nakashima K, Tran LS, Van Nguyen D, Fujita M, Maruyama K, Todaka D, Ito Y, Hayashi N, Shinozaki K, Yamaguchi-Shinozaki K. Functional analysis of a NAC-type transcription factor OsNAC6 involved in abiotic and biotic stress-responsive gene expression in rice. Plant J, 2007, 51: 617-630

[33]Kaneda T, Taga Y, Takai R, Iwano M, Matsui H, Takayama S, Isogai A, Che F S. The transcription factor OsNAC4 is a key positive regulator of plant hypersensitive cell death. EMBO J, 2009, 28: 926-936

[34]Park H C, Kim M L, Lee S M, Bahk J D, Yun D J, Lim C O, Hong J C, Lee S Y, Cho1 M J, Chung W S.Pathogen-induced binding of the soybean zinc finger homeodomain proteins GmZF-HD1 and GmZF-HD2 to two repeats of ATTA homeodomain binding site in the calmodulin isoform 4 (GmCaM4) promoter. Nucl Acid Res, 2007, 35: 1-12

[35]Wong H L, Sakamoto T, Kawasaki T, Umemura K, Shimamoto K. Down-regulation of metallothionein, a reactive oxygen scavenger, by the small GTPase OsRac1 in rice. Plant Physiol, 2004, 135: 1447-1456

[36]Ortega X, Velasquez J C, Perez L M. IP3 production in the hypersensitive response of lemon seeding against Alternaria alternata involves active protein tyrosine kinase but not a G protein. Biol Res, 2005, 38: 89-99

[37]Bargmann B O, Munnik T. The role of phospholipase D in plant stress responses. Curr Opin Plant Biol, 2006, 9: 515-522

[38]De Torres Zabela M, Fernandez-Delmond I, Niittyla T, Sanchez P, Grant M. Differential expression of genes encoding Arabidopsis phospholipases after challenge with virulent or avirulent Pseudomonas isolates. Mol Plant Microbe Interact, 2002, 15: 808-816

[39]Yamaguchi T, Kuroda M, Yamakawa H, Ashizawa T, Hirayae K, Kurimoto L, Shinya T, Shibuya N. Suppression of a phospholipase D gene, OsPLD, activates defense responses and increases disease resistance in rice. Plant Physiol, 2009, 150: 308-319
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