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Co-overexpression of Brassica juncea NPR1 (BjNPR1) and Trigonella foenum-graecum defensin (Tfgd) in transgenic peanut provides comprehensive but varied protection against Aspergillus flavus and Cercospora arachidicola

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Abstract

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Coexpression of two antifungal genes ( NPR1 and defensin ) in transgenic peanut results in the development of resistance to two major fungal pathogens, Aspergillus flavus and Cercospora arachidicola.

Abstract

Fungal diseases have been one of the principal causes of crop losses with no exception to peanut (Arachis hypogeae L.), a major oilseed crop in Asia and Africa. To address this problem, breeding for fungal disease resistance has been successful to some extent against specific pathogens. However, combating more than one fungal pathogen via breeding is a major limitation in peanut. In the present study, we demonstrated the potential use of co-overexpression of two genes, NPR1 and defensin isolated from Brassica juncea and Trigonella foenum-graecum respectively; that offered resistance towards Aspergillus flavus in peanut. The transgenic plants not only resisted the mycelial growth but also did not accumulate aflatoxin in the seeds. Resistance was also demonstrated against another pathogen, Cercospora arachidicola at varied levels; the transgenic plants showed both reduction in the number of spots and delay in the onset of disease. PCR, Southern and Western blot analysis confirmed stable integration and expression of the transgenes in the transgenic plants. The combinatorial use of the two pathogen resistance genes presents a novel approach to mitigate two important fungal pathogens of peanut.

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References

  • Arthur E, Waltking DW, Chan D, Dunn EJ, Humphries Kandler H, Sizoo E (2006) Liquid chromatographic analysis of aflatoxin using post-column photochemical derivatization: collaborative study. J AOAC Int 89:678–692

    Google Scholar 

  • Broekaert WF, Terras FR, Cammue BP, Osborn RW (1995) Plant defensins: novel antimicrobial peptides as components of the host defence system. Plant Physiol 108:1353–1358

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Broglie K, Chet I, Holliday M, Cressman R, Biddle P, Knowlton S, Mauvais CJ, Broglie R (1991) Transgenic plants with enhanced resistance to the fungal pathogen Rhizoctonia solani. Science 254:1194–1197

    Article  CAS  PubMed  Google Scholar 

  • Cao H, Li X, Dong X (1998) Generation of broad-spectrum disease resistance by overexpression of an essential regulatory gene in systemic acquired resistance. Proc Natl Acad Sci USA 95:6531–6536

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chern MS, Fitzgerald HA, Yadav RC, Canlas PE, Dong X, Ronald PC (2001) Evidence for a disease-resistance pathway in rice similar to the NPR1-mediated signaling pathway in Arabidopsis. Plant J 27:101–113

    Article  CAS  PubMed  Google Scholar 

  • Dana MM, Pintor-Toro JA, Cubero B (2006) Transgenic tobacco plants overexpressing chitinases of fungal origin show enhanced resistance to biotic and abiotic stress agents. Plant Physiol 142:722–730

    Article  PubMed Central  Google Scholar 

  • Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: version II. Plant Mol Biol Rep 1:19–21

    Article  CAS  Google Scholar 

  • Emani C, Garcia JM, Lopata-Finch E, Pozo MJ, Uribe P, Kim DJ, Sunilkumar G, Cook DR, Kenerley CM, Rathore KS (2003) Enhanced fungal resistance in transgenic cotton expressing an endochitinase gene from Trichoderma virens. Plant Biotechnol J 1:321–336

    Article  CAS  PubMed  Google Scholar 

  • Gaffney T, Friedrich L, Vernooij B, Negrotto D, Nye G, Uknes S, Ward E, Kessmann H, Ryals J (1993) Requirement of salicylic acid for the induction of systemic acquired resistance. Science 261:754–756

    Article  CAS  PubMed  Google Scholar 

  • Ghag SB, Shekhawat UKS, Ganapathi TR (2012) Petunia floral defensins with unique prodomains as novel candidates for development of Fusarium Wilt resistance in transgenic banana plants. PLoS One 7(6):e39557

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hajdukiewicz P, Svab Z, Maliga P (1994) The small, versatile pPZP family of Agrobacterium binary vectors for plant transformation. J Plant Mol Biol 25:989–994

    Article  CAS  Google Scholar 

  • Hoekema A, Hirsch PR, Hooykaas PJJ, Schilperoort RA (1983) A binary plant vector strategy based on separation of vir- and T-region of the Agrobacterium tumefaciens Ti-plasmid. Nature 303:179–180

    Article  CAS  Google Scholar 

  • Jha S, Chattoo BB (2010) Expression of a plant defensin in rice confers resistance to fungal phytopathogens. Transgenic Res 19:373–384

    Article  CAS  PubMed  Google Scholar 

  • Kumar V, Parkhi V, Kenerley CM, Rathore KS (2009a) Defence related gene expression and enzyme activities in transgenic cotton plants expressing an endochitinase gene from Trichoderma virens in response to interaction with Rhizoctonia solani. Planta 230:277–291

    Article  CAS  PubMed  Google Scholar 

  • Kumar AM, Sundaresha S, Rohini S (2009b) Resistance to Alternaria leaf spot disease in transgenic safflower (Carthamus tictorius L.) harboring a rice chitinase gene. Trans Plant J 3:113–118

    Google Scholar 

  • Kumar V, Joshi SG, Bell AA, Rathore KS (2013) Enhanced resistance against Thielaviopsis basicola in transgenic cotton plants expressing Arabidopsis NPR1 gene. Transgenic Res 22:359–368

    Article  PubMed  Google Scholar 

  • Lin WC, Lu CF, Wu JW, Cheng ML, Lin YM, Yang NS, Black L, Green SK, Wang JF, Cheng CP (2004) Transgenic tomato plants expressing the Arabidopsis AtNPR1 gene display enhanced resistance to a spectrum of fungal and bacterial diseases. Transgenic Res 13:567–581

    Article  CAS  PubMed  Google Scholar 

  • Makandar R, Essig JS, Schapaugh MA, Trick HN, Shah J (2006) Genetically engineered resistance to Fusarium head blight in wheat by expression of Arabidopsis NPR1. Mol Plant Microbe Interact 19:123–129

    Article  CAS  PubMed  Google Scholar 

  • Malnoy M, Jin Q, Borejsza-Wysocka EE, He SY, Aldwinckle HS (2007) Overexpression of the Apple MpNPR1 gene confers increased disease resistance in Malus domestica. Mol Plant Microbe Interact 20:1568–1580

    Article  CAS  PubMed  Google Scholar 

  • Meur G, Budatha M, Srinivasan T, Rajesh Kumar KR, Dutta Gupta A, Kirti PB (2008) Constitutive expression of Arabidopsis NPR1 confers enhanced resistance to the early instars of Spodoptera litura in transgenic tobacco. Physiol Plant 133:765–775

    Article  CAS  PubMed  Google Scholar 

  • Olli S, Kirti PB (2006) Cloning, characterization and antifungal activity of defensin Tfgd1 from Trigonella foenum-graecum L. J Biochem Mol Biol 39:278–283

    Article  CAS  PubMed  Google Scholar 

  • Osborn RW, De Samblanx GW, Thevissen K, Goderis I, Torrekens S, Van Leuven F, Attenborough S, Rees SB, Broekaert WF (1995) Isolation and characterisation of plant defensins from seeds of Asteraceae, Fabaceae, Hippocastanaceae and Saxifragaceae. FEBS Lett 368:257–262

    Article  CAS  PubMed  Google Scholar 

  • Parkhi V, Kumar V, Campbell LAM, Bell AA, Rathore KS (2010a) Expression of Arabidopsis NPR1 in transgenic cotton confers resistance to non-defoliating isolates of Verticillium dahliae but not the defoliating isolates. J Phytopathol 158:822–825

    Article  CAS  Google Scholar 

  • Parkhi V, Kumar V, Campbell LM, Bell AA, Shah J, Rathore KS (2010b) Resistance against various fungal pathogens and reniform nematode in transgenic cotton plants expressing Arabidopsis NPR1. Trans Res 19:959–975

    Article  CAS  Google Scholar 

  • Pieterse CM, Van Loon LC (2004) NPR1: the spider in the web of induced resistance signaling pathways. Curr Opin Plant Biol 7(4):456–464

    Article  CAS  PubMed  Google Scholar 

  • Punja ZK (2006) Recent developments toward achieving fungal disease resistance in transgenic plants. Can J Plant Pathol 28:S298–S308

    Article  CAS  Google Scholar 

  • Ramineni R, Sadumpati V, Khareedu VR, Vudem DR (2014) Transgenic pearl millet male fertility restorer line (ICMP451) and hybrid (ICMH451) expressing Brassica juncea nonexpressor of Pathogenesis Related Genes 1 (BjNPR1) exhibit resistance to downy mildew disease. PLoS One 9:e90839

    Article  PubMed  PubMed Central  Google Scholar 

  • Rivero M, Furman N, Mencacci N, Picca P, Toum L, Lentz E, Bravo-Almonacid F, Mentaberry A (2012) Stacking of antimicrobial genes in potato transgenic plants confers increased resistance to bacterial and fungal pathogens. J Biotechnol 157:334–343

    Article  CAS  PubMed  Google Scholar 

  • Rohini VK, Rao KS (2001) Transformation of peanut (Arachi hypogaea L.) with tobacco chitinase gene: variable responses of transformants to leaf spot disease. Plant Sci 16:889–898

    Article  Google Scholar 

  • Ryals JA, Neuenschwander UH, Willits MG, Molina A, Steiner HY, Hunt MD (1996) Systemic acquired resistance. Plant Cell 8:1809–1819

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sadumpati V, Kalamburb M, Vudema DR, Kirti PB, Kharredu VR (2013) Transgenic indica rice lines, expressing Brassica juncea nonexpressor of pathogenesis-related genes 1 (BjNPR1), exhibit enhanced resistance to major pathogens. J Biotechnol 166:114–121

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: laboratory manual, 2nd edn. Coldspring Harbor Press, New York

    Google Scholar 

  • Sundaresha S, Kumar MA, Rohini S, Math SA, Keshamma E, Chandrashekar SC, Udayakumar M (2010) Enhanced protection against two major fungal pathogens of peanut, Cercospora arachidicola and Aspergillus flavus in transgenic peanut over-expressing a tobacco β 1–3 glucanase. Eur J Plant Pathol 126:497–508

    Article  CAS  Google Scholar 

  • Thevissen K, Warnecke DC, Francois IEJA, Leipelt M, Heinz E, Ott C, Zähringer U, Thomma BPHJ, Ferket KKA, Cammue BPA (2004) Defensins from insects and plants interact with fungal glucosylceramides. J Biol Chem 279:3900–3905

    Article  CAS  PubMed  Google Scholar 

  • Thirulogachandar V, Prachi P, Vaishnavi CS, Reddy MK (2011) An affinity-based genome walking method to find transgene integration loci in transgenic genome. Anal Biochem 416:196–201

    Article  CAS  PubMed  Google Scholar 

  • Thomma BPHJ, Cammue BPA, Thevissen K (2002) Plant defensins. Planta 216:193–202

    Article  CAS  PubMed  Google Scholar 

  • Vasavirama K, Kirti PB (2013) Constitutive expression of a fusion gene comprising Trigonella foenum-graecum defensin (Tfgd2) and Raphanus sativus antifungal protein (RsAFP2) confers enhanced disease and insect resistance in transgenic tobacco. Plant Cell Tissue Organ Cult 115:309–319

    Article  CAS  Google Scholar 

  • Verma SS, Yajima WR, Rahman MH, Shah S, Liu JJ, Ekramoddoullah AK, Kav NN (2012) A cysteine-rich antimicrobial peptide from Pinus monticola (PmAMP1) confers resistance to multiple fungal pathogens in canola (Brassica napus). Plant Mol Biol 79:61–74

    Article  CAS  PubMed  Google Scholar 

  • Vijayan S, Kirti PB (2012) Mungbean plants expressing BjNPR1 exhibit enhanced resistance against the seedling rot pathogen, Rhizoctonia solani. Transgenic Res 21:193–200

    Article  CAS  PubMed  Google Scholar 

  • Wally O, Jayaraj J, Punja ZK (2009) Broad-spectrum disease resistance to necrotrophic and biotrophic pathogens in transgenic carrots (Daucus carota L.) expressing an Arabidopsis NPR1 gene. Planta 231:131–141

    Article  CAS  PubMed  Google Scholar 

  • Zhang X, Francis MI, Dawson WO, Graham JH, Orbović V, Triplett EW, Mou Z (2010) Over-expression of the Arabidopsis NPR1 gene in citrus increases resistance to citrus canker. Eur J Plant Pathol 128:91–100

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors wish to acknowledge financial support from NABARD (National Bank for Agriculture and Rural Development), India and Biotech Hub, Department of Biotechnology, India.

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    Authors and Affiliations

    1. Department of Crop Physiology, University of Agricultural Sciences, GKVK, Bengaluru, India

      S. Sundaresha, Sreevathsa Rohini, V. K. Appanna, Manoj-Kumar Arthikala, N. B. Shanmugam, N. B. Shashibhushan, C. M. Hari Kishore & M. Udayakumar

    2. Department of Pathology, University of Agricultural Sciences, GKVK, Bengaluru, India

      S. Sundaresha

    3. Department of Plant Sciences, School of Life Science, University of Hyderabad, Hyderabad, India

      P. B. Kirti

    4. ICAR-Central Potato Research Institute, Shimla, HP, India

      S. Sundaresha

    5. ICAR-National Research Centre on Plant Biotechnology, LBS Centre, Pusa Campus, New Delhi, India

      Sreevathsa Rohini

    6. Escuela Nacional de Estudios Superiores, Universidad Nacional Autónoma de México (UNAM), León, 37684, Guanajuato, Mexico

      Manoj-Kumar Arthikala

    7. Department of Botany, Annamalai University, Annamalai Nagar, Chidambaram, India

      V. K. Appanna & R. Pannerselvam

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    1. S. Sundaresha
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    4. Manoj-Kumar Arthikala
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    5. N. B. Shanmugam
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    9. P. B. Kirti
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    10. M. Udayakumar
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    Correspondence to M. Udayakumar.

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    Communicated by M. Prasad.

    S. Sundaresha, S. Rohini and V. K. Appanna have made equal contributions to the work.

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    Sundaresha, S., Rohini, S., Appanna, V.K. et al. Co-overexpression of Brassica juncea NPR1 (BjNPR1) and Trigonella foenum-graecum defensin (Tfgd) in transgenic peanut provides comprehensive but varied protection against Aspergillus flavus and Cercospora arachidicola . Plant Cell Rep 35, 1189–1203 (2016). https://doi.org/10.1007/s00299-016-1945-7

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