Project description:Trichoderma harzianum CECT 2413 expression in liquid basal medium and in the presence of glucose, chitin or tomato plants. Four different experimental conditions were carried out: basal (MS), glucose (MS-G), chitin (MS-Q) and tomato plant (MS-P). Two biological replicates were analyzed by microarray for each experimental condition. Three independent cultures of mycelium were pooled for each biological replicate.
Project description:A self-designed Trichoderma high density oligonuclotide (HDO) microarray (Roche-NimbleGen, Inc., Madison, WI, USA) was constructed in a similar way than a previous Trichoderma HDO microarray (Samolski et al., 2009). The microarray was composed of 392,779 60-mer probes designed against 13,443 EST-derived transcripts (Trichochip-1) and the genomes of T. atroviride (11,100 genes) and T. virens (11,643 genes). The Trichochip-1 ESTs were obtained from 28 cDNA libraries from eight different species (representing the biodiversity of this genus: T. harzianum, T. atroviride, T. asperellum, T. viride, T. longibrachiatum, T. virens, T. stromaticum and T. aggresivum), under a wide range of growth conditions, including biocontrol-related conditions and different nutritional situations (VizcaÃno et al., 2006). The Trichochip1 EST database was generated in the TrichoEST project funded by the EU (QLK3-CT-2002-02032). Confrontations were carried out between the T. harzianum T34 or nox1 overexpressed transformant Tnox5 and P. ultimum. Agar plugs cut from the growing edge of a 4-day colony of Trichoderma and Pythium were placed 2 cm from the border on the opposite side of the same petri plates containing PDA covered with sterile cellophane sheets. Dual cultures were allowed to grow at 25 ºC and mycelia were collected from the interaction zone in confrontations between P. ultimum and T. harzianum T34 or Tnox5 strains. Seven PDA plates were used for each condition considered and RNAs were extracted and the corresponding cDNAs were use to hybridize by triplicate the Trichoderma HDO microarray.
Project description:The plant recognition specific PCA cluster mediates early chemical communication between plant and fungus, is required for colonization and it is likely responsible for the high potential of T. harzianum and closely related species for biocontrol applications.
Project description:The regulation of plant biomass degradation by fungi is critical to the carbon cycle, and applications in bioproducts and biocontrol. Trichoderma harzianum is an important plant biomass degrader, enzyme producer, and biocontrol agent, but few putative major transcriptional regulators have been deleted in this species. The T. harzianum ortholog of the transcriptional activator XYR1/XlnR/XLR-1 was deleted, and the mutant strains were analyzed by growth profiling, enzymatic activities, and transcriptomics on cellulose. From plate cultures, the Δxyr1 mutant had reduced growth on D-xylose, xylan, and cellulose, and from shake-flask cultures with cellulose, the Δxyr1 mutant had ~ 90% lower β-glucosidase activity, and no detectable β-xylosidase or cellulase activity. The comparison of the transcriptomes from 18 h shake-flask cultures on D-fructose, without a carbon source, and cellulose, showed major effects of XYR1 deletion whereby the Δxyr1 mutant on cellulose was transcriptionally most similar to the cultures without a carbon source. The cellulose induced 43 plant biomass-degrading CAZymes including xylanases as well as cellulases, and most of these had massively lower expression in the Δxyr1 mutant. Expression of a sub-set of carbon catabolic enzymes, other transcription factors, and sugar transporters was also lower in the Δxyr1 mutant on cellulose. In summary, T. harzianum XYR1 is the master regulator of cellulases and xylanases, as well as regulating carbon catabolic enzymes.
