Trichoderma Transformation Methods

  • Mónica G. Malmierca
  • Rosa E. Cardoza
  • Santiago GutiérrezEmail author
Part of the Fungal Biology book series (FUNGBIO)


Trichoderma includes a huge variety of fungal species with an increasing interest from several points of view. Many of their species produce primary or secondary metabolites with importance in pharmaceutical industry or in the biocontrol of significant phytopathogenic fungi (Cardoza et al., Curr Genet 34:50–59, 2005; Sivasithamparam and Ghisalberti, Trichoderma and Gliocladium, London, pp 139–191). Other Trichoderma species produce enzymes with a remarkable industrial importance and even other strains from Trichoderma brevicompactum (Tijerino et al., Fungal Genet Biol 48:285–296, 2011a, Toxins (Basel) 3:1220–1232, 2011b) or Trichoderma longibrachiatum (Alanio et al., Clin Infect Dis 46:e116–118, 2008) have potential pathogenic activity against plants and animals, including human beings, respectively. The practical applications of this genus have resulted in an increasing interest in the development of efficient transformation procedures, in order to select transgenic strains with valuable phenotypes. These procedures should be essential tools to characterize the physiological roles of the increasing number of genes available as a result of the fungal genomic projects ( (Grigoriev et al., Nucleic Acids Res 40:D26–32, 2012).


Trichoderma Phytopathogenic fungi Fungi transformation techniques Electroporation Fungal genomic projects Protoplast transformation 



 We thank Dr. Elías R. Olivera for constructive comments and critical reading of the manuscript. Dr. Gutiérrez receives grant-aided support from the Ministry of Science and Innovation of Spain (AGL2012-40041-C02-02) and from the Junta de Castilla y León (LE125A12-2).


  1. Alanio A, Brethon B, Feuilhade de Chauvin M, Kerviler E, Leblanc T, Lacorix C, Baruchel A, Menotti J (2008) Invasive pulmonary infection due to Trichoderma longibrachiatum mimicking invasive aspergillosis in a neutropenic patient successfully treated with voriconazole combined with caspofungin. Clin Infect Dis 46: e116–e118PubMedCrossRefGoogle Scholar
  2. Cardoza RE, Moralejo FJ, Gutiérrez S, Casqueiro J, Fierro F, Martín JF (1998) Characterization and nitrogen source regulation at the transcriptional level of the gdhA gene of Aspergillus awamori encoding an NADP-dependent glutamate dehydrogenase. Curr Genet 34:50–59PubMedCrossRefGoogle Scholar
  3. Cardoza RE, Hermosa MR, Vizcaíno JA, Sanz L, Monte E, Gutiérrez S (2005) Secondary metabolites produced by Trichoderma and their importance in the biocontrol process. In: Mellado E, Barredo JL (eds) Microorganisms for industrial enzymes and biocontrol. Research Signpost, pp 1–22. ISBN: 81-308-0040-3Google Scholar
  4. Cardoza RE, Vizcaíno JA, Hermosa R, Monte E, Gutiérrez S (2006) A comparison of the phenotypic and genetic stability of recombinant Trichoderma spp. generated by protoplast- and Agrobacterium-mediated transformation. J Microbiol 44:383–395PubMedGoogle Scholar
  5. Cardoza RE, Hermosa MR, Vizcaíno JA, González FJ, Llobell A, Monte E, Gutiérrez S (2007) Partial silencing of a hydroxy-methylglutaryl-CoA reductase encoding gene in Trichoderma harzianum CECT 2413 results in a lower level of resistance to lovastatin and a lower antifungal activity. Fungal Genet Biol 44: 269–283PubMedCrossRefGoogle Scholar
  6. De Groot MJA, Bundock P, Hooykaas PJJ, Beijersbergen AGM (1998) Agrobacterium tumefaciens-mediated transformation of filamentous fungi. Nat Biotechnol 16:839–842PubMedCrossRefGoogle Scholar
  7. Grigoriev IV, Nordberg H, Shabalov I, Aerts A, Cantor M, Goodstein D, Kuo A, Minovitsky S, Nikitin R, Ohm RA, Otillar R, Poliakov A, Ratnere I, Riley R, Smirnova T, Rokhsar D, Dubchak I (2012) The genome portal of the Department of Energy Joint Genome Institute. Nucleic Acids Res 40:D26–D32PubMedCrossRefPubMedCentralGoogle Scholar
  8. Gruber F, Visser J, Kubicek CP, de Graaff LH (1990) The development of a heterologous transformation system for the cellulolytic fungus Trichoderma reesei based on a pyrG-negative mutant strain. Curr Genet 18: 71–76PubMedCrossRefGoogle Scholar
  9. Gruber F, Bicker W, Oskolkova OV, Tschachler E, Bochkov VN (2012) A simplified procedure for semi-targeted lipidomic analysis of oxidized phosphatidylcholines induced by UVA irradiation. J Lipid Res 53:1232–1242PubMedCrossRefPubMedCentralGoogle Scholar
  10. Guangtao Z, Seiboth B, Wen C, Yaohua Z, Xian L, Wang T (2010) A novel carbon source-dependent genetic transformation system for the versatile cell factory Hypocrea jecorina (anamorph Trichoderma reesei). FEMS Microbiol Lett 303:26–32PubMedCrossRefGoogle Scholar
  11. Hooykas PJJ, Roobol C, Schilperoort RA (1979) Regulation of the transfer of Ti-plasmids of Agrobacterium tumefaciens. J Gen Microbiol 110: 99–109CrossRefGoogle Scholar
  12. Lorito M, Hayes CK, Di Pietro A, Harman GE (1993) Biolistic transformation of Trichoderma harzianum and Gliocladium virens using plasmid and genomic DNA. Curr Genet 24:349–356PubMedCrossRefGoogle Scholar
  13. Mach RL, Schindler M, Kubicek CP (1994) Transformation of Trichoderma reesei based on hygromycin B resistance using homologous expression signals. Curr Genet 25:567–570PubMedCrossRefGoogle Scholar
  14. Magaña-Ortíz D, Coconi-Linares N, Ortíz-Vazquez E, Fernández F, Loske AM, Gómez-Lim MA (2013) A novel and highly efficient method for genetic transformation of fungi employing shock waves. Fungal Genet Biol 56:9–16PubMedCrossRefGoogle Scholar
  15. Malmierca MG, Cardoza RE, Alexander NJ, McCormick SP, Hermosa R, Monte E, Gutiérrez S (2012) Involvement of Trichoderma trichothecenes in the biocontrol activity and induction of plant defense-related genes. Appl Environ Microbiol 78:4856–4868PubMedCrossRefPubMedCentralGoogle Scholar
  16. Malmierca MG, Cardoza RE, Alexander NJ, McCormick SP, Collado IG, Hermosa R, Monte E, Gutiérrez S (2013) Relevance of trichothecenes in fungal physiology: disruption of tri5 in Trichoderma arundinaceum. Fungal Genet Biol 53:22–33PubMedCrossRefGoogle Scholar
  17. Mozo T, Hooykaas PJ (1991) Electroporation of megaplasmids into Agrobacterium. Plant Mol Biol 16: 917–918PubMedCrossRefGoogle Scholar
  18. Penttila M, Nevalainen H, Ratto M, Salminen E, Knowles J (1987) A versatile transformation system for the cellulotytic filamentous fungus Trichoderma reesei. Gene 61:155–164PubMedCrossRefGoogle Scholar
  19. Punt PJ, Oliver RP, Dingemanse MA, Pouwels PH, van den Hondel CAMJJ (1987) Transformation of Aspergillus based on hygromycin B resistance marker from Escherichia coli. Gene 56:117–124PubMedCrossRefGoogle Scholar
  20. Sánchez-Torres P, González R, Pérez-González JA, Gozález-Candelas L, Ramón D (1994) Development of a transformation system for Trichoderma longibrachiatum and its use for constructing multicopy transformants for the egl1 gene. Appl Microbiol Biotechnol 41:440–446PubMedGoogle Scholar
  21. Schuster A, Bruno KS, Collett JR, Baker SE, Selboth B, Kubicek CP, Schmoll M (2012) A versatile toolkit for high throughput functional genomics with Trichoderma reesei. Biotechnol Biofuels 5:1PubMedCrossRefPubMedCentralGoogle Scholar
  22. Sivan A, Stasz TE, Hemmat M, Hayes CK, Harman GE (1992) Transformation of Trichoderma spp. with plasmids conferring hygromycin B resistance. Mycologia 84:687–694CrossRefGoogle Scholar
  23. Sivasithamparam K, Ghisalberti EL (1998) Secondary metabolism in Trichoderma and Gliocladium. In: Harman GE, Kubicek CP (eds) Trichoderma and gliocladium, vol 1. Taylor and Francis, London, pp 139–191Google Scholar
  24. Te’o VS, Bergquist PL, Nevalainen KM (2002) Biolisitic transformation of Trichoderma reesei using the Bio-Rad seven barrels hepta adaptor system. J Microbiol Methods 51:393–399PubMedCrossRefGoogle Scholar
  25. Tijerino A, Cardoza RE, Moraga J, Malmierca MG, Vicente F, Aleu J, Collado IG, Gutiérrez S, Monte E, Hermosa R (2011a) Overexpression of the trichodiene synthase gene tri5 increases trichodermin production and antimicrobial activity in Trichoderma brevicompactum. Fungal Genet Biol 48:285–296PubMedCrossRefGoogle Scholar
  26. Tijerino A, Hermosa R, Cardoza RE, Moraga J, Malmierca MG, Aleu J, Collado IG, Monte E, Gutierrez S (2011b) Overexpression of the Trichoderma brevicompactum tri5 gene: effect on the expression of the trichodermin biosynthetic genes and on tomato seedlings. Toxins (Basel) 3:1220–1232CrossRefGoogle Scholar
  27. Yang L, Yang Q, Sun K, Tian Y (2011) Agrobacterium tumefaciens of ChiV gene to Trichoderma harzianum. Appl Biochem Biotechnol 163:937–945PubMedCrossRefGoogle Scholar
  28. Zhong YH, Xiao LW, Wang TH, Jiang Q (2007) Agrobacterium-mediated transformation (AMT) of Trichoderma reesei as an efficient tool for random insertional mutagenesis. Appl Microbiol Biotechnol 73:1348–1354PubMedCrossRefGoogle Scholar
  29. Zhu T, Wang W, Yang X, Wang K, Zhifeng C (2009) Construction of two gateway vectors for gene expression in fungi. Plasmid 62:128–133PubMedCrossRefGoogle Scholar

Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  • Mónica G. Malmierca
    • 1
  • Rosa E. Cardoza
    • 1
  • Santiago Gutiérrez
    • 1
    Email author
  1. 1.Department of Molecular BiologyUniversity of LeónPonferradaSpain

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