Advertisement

Targeting Trichothecene Biosynthetic Genes

  • Songhong Wei
  • Theo van der Lee
  • Els Verstappen
  • Marga van Gent
  • Cees WaalwijkEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1542)

Abstract

Biosynthesis of trichothecenes requires the involvement of at least 15 genes, most of which have been targeted for PCR. Qualitative PCRs are used to assign chemotypes to individual isolates, e.g., the capacity to produce type A and/or type B trichothecenes. Many regions in the core cluster (consisting of 12 genes) including intergenic regions have been used as targets for PCR, but the most robust assays are targeted to the tri3 and tri12 genes. Quantitative PCRs, that work across trichothecene-producing members of the Fusarium head blight complex, are described along with procedures to quantify the amount of fungal biomass in wheat samples. These assays are directed to the chemotype(s) present in field samples and quantify the total fungal biomass of trichothecene-producing fungi, irrespective of their genetic identity.

Key words

Trichothecenes PCR Quantitative PCR 

Notes

Acknowledgments

This work was supported by the MycoRed project (FP7 Food Quality and Safety Priority—Large Collaborative Project—GA 222690) and the Dutch Main Board for Arable Products.

References

  1. 1.
    Alexander NJ, Proctor RH, McCormick SP (2009) Genes, gene clusters, and biosynthesis of trichothecenes and fumonisins in Fusarium. Toxin Rev 28:198–215CrossRefGoogle Scholar
  2. 2.
    Kimura M, Tokai T, Takahashi-Ando N et al (2007) Molecular and genetic studies of fusarium trichothecene biosynthesis: pathways, genes, and evolution. Biosci Biotechnol Biochem 71:2105–2123CrossRefPubMedGoogle Scholar
  3. 3.
    McCormick SP, Stanley AM, Stover NA et al (2011) Trichothecenes: from simple to complex mycotoxins. Toxins 3:802–814CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Proctor RH, McCormick SP, Alexander NJ et al (2009) Evidence that a secondary metabolic biosynthetic gene cluster has grown by gene relocation during evolution of the filamentous fungus Fusarium. Mol Microbiol 74:1128–1142CrossRefPubMedGoogle Scholar
  5. 5.
    Rep M, Kistler HC (2010) The genomic organization of plant pathogenicity in Fusarium species. Curr Opin Plant Biol 13:420–426CrossRefPubMedGoogle Scholar
  6. 6.
    Alexander NJ, McCormick SP, Waalwijk C et al (2011) The Genetic Basis for 3-ADON and 15-ADON Trichothecene Chemotypes in Fusarium. Fungal Genet Biol 48:485–495CrossRefPubMedGoogle Scholar
  7. 7.
    Brown DW, McCormick SP, Alexander NJ et al (2001) A Genetic and Biochemical Approach to Study Trichothecene Diversity in Fusarium sporotrichioides and Fusarium graminearum. Fungal Genet Biol 32:121–133CrossRefPubMedGoogle Scholar
  8. 8.
    Brown DW, McCormick SP, Alexander NJ et al (2002) Inactivation of a cytochrome P-450 is a determinant of trichothecene diversity in Fusarium species. Fungal Genet Biol 36:224–233CrossRefPubMedGoogle Scholar
  9. 9.
    Lee T, Oh DW, Kim HS et al (2001) Identification of deoxynivalenol- and nivalenol-producing chemotypes of Gibberellazeae by using PCR. Appl Environ Microbiol 67:2966–2972CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Zhang H, Zhang Z, van der Lee T et al (2010) Population genetic analyses of Fusarium asiaticum populations from barley suggest a recent shift favoring 3ADON producers in southern China. Phytopathology 100:328–336CrossRefPubMedGoogle Scholar
  11. 11.
    Ward TJ, Bielawski JP, Kistler HC et al (2002) Ancestral polymorphism and adaptive evolution in the trichothecene mycotoxin gene cluster of phytopathogenic Fusarium. Proc Natl Acad Sci U S A 99:9278–9283CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Waalwijk C, Köhl J, De Vries I et al (2009) Fusarium in winter tarwe (in 2007 en 2008) (in Dutch). Wageningen, The Netherlands, Plant Research International. Report 272Google Scholar
  13. 13.
    Kulik T (2011) Development of TaqMan assays for 3ADON, 15ADON and NIV Fusarium genotypes based on Tri12 gene. Cereal Res Commun 39:200–214CrossRefGoogle Scholar
  14. 14.
    O’Donnell K, Kistler HC, Tacke BK et al (2000) Gene genealogies reveal global phylogeographic structure and reproductive isolation among lineages of Fusarium graminearum, the fungus causing wheat scab. Proc Natl Acad Sci U S A 97:7905–7910CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Yang LJ, van der Lee TAJ, Yang XJ et al (2008) Fusarium Populations on Chinese Barley Show a Dramatic Gradient in Mycotoxin Profiles. Phytopathology 98:719–727CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Songhong Wei
    • 1
  • Theo van der Lee
    • 2
  • Els Verstappen
    • 2
  • Marga van Gent
    • 2
  • Cees Waalwijk
    • 2
    Email author
  1. 1.College of Plant ProtectionShenyang Agricultural UniversityShenyangPeople’s Republic of China
  2. 2.Biointeractions and Plant HealthWageningen URWageningenThe Netherlands

Personalised recommendations