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作物学报 ›› 2012, Vol. 38 ›› Issue (06): 980-987.doi: 10.3724/SP.J.1006.2012.00980

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

水稻稻瘟病菌诱导表达启动子OsQ16p的克隆与功能分析

王光，吴智丹，张磊，刘凤权，邵敏*   

  1. 南京农业大学植物保护学院/农作物生物灾害综合治理教育部重点实验室，江苏南京210095
  • 收稿日期:2011-12-22 修回日期:2012-02-22 出版日期:2012-06-12 网络出版日期:2012-03-29
  • 通讯作者: 邵敏, E-mail: minshao@njau.edu.cn
  • 基金资助:

    本研究由国家高技术研究发展计划(863计划)项目(2007AA10Z188)和国家转基因生物新品种培育项目(2009ZX08001-005B)资助。

Cloning and Functional Analysis of Magnaporthe oryzae-Induced Promoter OsQ16p in Rice

WANG Guang,WU Zhi-Dan,ZHANG Lei,LIU Feng-Quan,SHAO Min*   

  1. College of Plant Protection, Nanjing Agricultural University/Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing 210095, China
  • Received:2011-12-22 Revised:2012-02-22 Published:2012-06-12 Published online:2012-03-29
  • Contact: 邵敏, E-mail: minshao@njau.edu.cn

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被引次数

10

摘要： qRT-PCR分析表明，日本晴OsQ16基因受稻瘟病菌诱导表达。利用PCR技术从日本晴基因组中克隆了该基因编码区5′端上游1 229 bp的启动子序列，命名为OsQ16p。用其取代pBI121中gus基因上游的CaMV35S启动子，构建重组表达载体pBIQ16p，经农杆菌介导转化日本晴，获得转基因植株。GUS组织化学染色和qRT-PCR分析表明: (1) gus基因在抗性愈伤组织和阳性转基因植株中均能表达; (2)转基因植株在接种稻瘟病菌后12 h，GUS表达量是处理前的2.7倍; (3)抗病相关信号分子水杨酸(salicylic acid，SA)和茉莉酸甲酯(methyl jasmonate，MeJA)喷施转基因植株叶面后12 h，GUS表达量分别为处理前的3.1倍和3.5倍。以上结果表明，OsQ16p启动子具有启动活性，并明显受稻瘟病菌、MeJA和SA诱导表达。

关键词: 转基因水稻, 诱导型启动子, OsQ16p, GUS, 稻瘟病菌

Abstract: The qRT-PCR analysis showed that, in Nipponbare (Oryza sativa L. ssp japonica), the expressionof OsQ16 gene was induced by Magnaporthe grisea. The 1 299 bp-fragment of 5'-end of OsQ16 gene, named as OsQ16p,was amplified by PCR from Nipponbare. The plasmids pBIQ16p was constructed by replacing the CaMV35S promoter of pBI121 with the OsQ16p, and transformed into Nipponbare through Agrobacterium-mediated transformation. The analysis of GUS activity and qRT-PCR showed that gus gene could express in transgenic plants and callui. The expression of gus gene in transgenic plants was obviously enhanced by M. grisea. After treatment with the resistance-related signaling molecules SA and MeJA, the GUS activities in transgenic plants were increased by 3.1- and 3.5-fold, respectively. It suggested that M. grisea, SA and MeJA were inductive factors of OsQ16p promoter.

Key words: Transgenic rice, Inducible promoter, OsQ16p, GUS, Magnaporthe grisea

[1]Nie L-N(聂丽娜), Xia L-Q(夏兰琴), Xu Z-S(徐兆师). Progress on cloning and functional study of plant gene promoters. J Plant Genet Resour (植物遗传资源学报), 2008, 9(3): 385–391 (in Chinese with English abstract)

[2]Yin Y, Chen L, Beachy R. Promoter elements required for phloem-specific gene expression from the RTBV promoter in rice. Plant J, 1997, 12: 1179–1188

[3]Nitz I, Berkefeld H, Puzio P S, Grundler F M. Pyk10, a seedling and root specific gene and promoter from Arabidopsis thaliana. Plant Sci, 2001, 161: 337–346

[4]Lu J(路静), Zhao H-Y(赵华燕), He Y-K(何奕昆), Song Y-R(宋艳茹). Advances in the study and application of higher plant promoter. Prog Nat Sci (自然科学进展), 2004, 14(8): 856–862 (in Chinese)

[5]Sunilkumar G, Mohr L, Lopata-Finch E. Developmental and tissue-specific expression of CaMV 35S promoter in cotton as revealed by GFP. Plant Mol Biol, 2002, 50: 463–474

[6]Chatterjee M, Banerjee A K, Hannapel D J. A BELL1-like gene of potato is light activated and wound inducible. Plant Physiol, 2007, 145: 1435–1443

[7]Cai Y-R(柴玉荣), Tian F(田芳), Liu H-T(刘红涛). Cloning and functional analysis of Dunaliella salina promoter rbcS. China Biotechnol (中国生物工程杂志), 2008, 28(4): 47–52 (in Chinese)

[8]Zhu Q, Song B T, Zhang C, Liu J. Construction and functional characteristics of tuber-specific and cold-inducible chimeric promoters in potato. Plant Cell Rep, 2008, 27: 47–55

[9]Narusaka Y, Nakashima K, Shinwari Z K, Sakuma Y, Furihata T, Abe H, Narusaka M, Shinozaki K, Yamaguchi-Shinozaki K. Interaction between two cis-acting elements, ABRE and DRE, in ABA-dependent expression of Arabidopsis rd29A gene in response to dehydration and high-salinity stresses. Plant J, 2003, 34: 137–148

[10]Shen Q, Chen C N, Brands A, Pan S M, Ho T H. The stress- and abscisic acid-induced barley gene HVA22: developmental regulation and homologues in diverse organisms. Plant Mol Biol, 2001, 45: 327–340

[11]Li A, Chen L L, Ren H Y, Wang X C, Huang R F. Analysis of the essential DNA region for OsEBP-89 promoter in response to methyl jasmonic acid. Sci China (Ser C: Life Sci), 2008, 51(3): 280–285

[12]Li N(李宁), Fan S-J(樊守金), Zhang Z-Y(张增艳). Progress in plant promoters related to disease resistance. J Plant Genet Resour (植物遗传资源学报), 2007, 8(2): 234–239 (in Chinese with English abstract)

[13]Baker B, Zambryski P, Staskawicz B, Dinesh-Kumar S P. Signaling in plant-microbe interactions. Science, 1997, 276: 726–728

[14]Lescot M, Déhais P, Thijs G, Marchal K, Moreau Y, Van de Peer Y, Rouzé P, Rombauts S. PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences. Nucl Acids Res, 2002, 30: 325–327

[15]Sambrook J, Fritsch E F, Maniatis T. Molecular Cloning: a Laboratory Manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press, 1989

[16]Huang J Q, Wei Z M, An H L, Zhu Y X. Agrobacterium tumefaciens-mediated transformation of rice with the spider insecticidal gene conferring resistance to leaffolder and striped stem borer. Cell Res, 2001, 11: 149–155

[17]Hiei Y, Ohta S, Komari T, Kumashiro T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J, 1994, 6: 271–282

[18]Jefferson R A, Kavanagh T A, Bevan M W. GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J, 1987, 6: 3901–3907

[19]Zhang Z-H(张智慧), Nie Y-F(聂燕芳), He L(何磊), Li Y-F(李云锋), Wang Z-Z(王振中). Resistance-related defense enzymes and endogenous salicylic acid induced by exogenous methyl jasmonate in rice against blast disease. Acta Phytopathol Sin (植物病理学报), 2010, 40(4): 395–403 (in Chinese with English abstract)

[20]Lee M W, Qi M, Yang Y N. A novel jasmonic acid-inducible rice myb gene associates with fungal infection and host cell death. Mol Plant Microbe Interact, 2001, 14: 527–535

[21]Wu G-Q(吴功庆), Li Y-N(李亚男), Chen D-Q(陈大清). Anaerobic gene expression and regulation mechanism in plants. J Trop Subtrop Bot (热带亚热带植物学报), 2006, 14(1): 87–92 (in Chinese with English abstract)

[22]Fukuda Y. Interaction of tobacoo nuclear protein with an elicitor-resistance element in the promoter of a basic class I chitinase gene. Plant Mol Biol, 1997, 34: 81–87

[23]Balbi V, Devoto A. Jasmonate signaling network in Arbidopsis thaliana: crucial regulatory nodes and new physiological scenarios. New Phytol, 2008, 177: 301–318

[24]Sa Q, Wang Y, Li W, Zhang L, Sun Y. The promoter of an antifungal protein gene from Gastrodia elata confers tissue-specific and fungus-inducible expression patterns and responds to both salicylic acid and jasmonic acid. Plant Cell Rep, 2003, 22: 79–84

[25]Moreno A B, Penas G, Rufat M, Bravo J M, Estopa M, Messeguer J, San Segundo B. Pathogen-induced production of the antifungal AFP protein from A spergillus giganteus confers resistance to the blast fungus Magnapor the grisea in transgenic rice. Mol Plant Microbe Interact, 2005, 18: 960–972

[26]Choi J J, Klosterma S J, Hadwiger L A. A promoter from pea gene DRR206 is suitable to regulate an elicitor-coding gene and develop disease resistance. Phytopathology, 2004, 94: 651–660

[27]Durrant W E, Dong X. Systematic acquired resistance. Annu Rev Phytopathol, 2004, 42: 185–209

[28]Halim V A, Vess A, Scheel D, Rosahl S. The role of salicylic acid and jasmonic acid in pathogen defence. Plant Biol, 2006, 8: 307–313

[29]Robert-Seilanizatz A, Nacarro L, Bari R, Jones J D G. Pathological hormone imbalances. Curr Opin Plant Biol, 2007, 10: 372–379

[30]Wu G-Z(吴国昭), Xie L-J(谢丽君), Song Y-Y(宋圆圆), Chen M(陈敏), Zeng R-S(曾任森). The physiological and biochemical mechanisms of disease resistance of Oryza rufipogon Griff. indigenous to Gaozhou, Guangdong against Pyricularia gisea induced by exogenously applied plant signal compounds. Acta Agric Boreali-Occident Sin (西北农业学报), 2009, 18(3): 254–258 (in Chinese with English abstract)

[31]Peng X-X(彭喜旭), Hu Y-J(胡耀军), Tang X-K(唐新科), Zhou P-L(周平兰), Deng X-B(邓小波), Wang H-H(王海华). Isolation and expression profiles of rice WRKY30 induced by jasmonic acid application and fungal pathogen infection. Sci Agric Sin (中国农业科学), 2011, 44(12): 2454–2461 (in Chinese with English abstract)
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