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:Compared with other organisms or conventional synthesis methods, the biogenic synthesis of nanoparticles from fungi has multiple advantages. Although the process is simple, nontoxic, and inexpensive, the synthesis mechanism remains largely undetermined. In this research, we focused on the proteins present in the aqueous filtrate of a native strain of Trichoderma harzianum, which is used as a reducing and stabilizing medium in the biogenic synthesis of zinc oxide nanoparticles (ZnONPs) and iron oxide nanoparticles (FeONPs). SDS‒PAGE analysis revealed distinct protein profiles in the culture medium (CM) and aqueous filtrate (AF), identifying through proteomic approach approximately 99 proteins in the CM and 304 in the AF, with 91% of the proteins only present in AF. From AF functional annotation classified the proteins mainly involved in metabolic processes and stress responses, with a notable presence of oxidoreductases. Successfully, ZnO and FeO NPs were synthesized from the aqueous filtrate and which presented a bouquet or irregular morphology. Characterization by TEM and EDS confirmed the successful synthesis of ZnONPs with an average size of 660 ± 109 nm and FeONPs with an average size of 18 ± 7 nm. Dynamic light scattering (DLS) analysis indicated that the ZnONPs presented low size variability and good stability, whereas the FeONPs presented a bimodal size distribution. Both NPs have thermal stability at 400 ºC meanwhile X-ray diffraction (XRD) confirmed the crystalline nature of both types of nanoparticles. FTIR analysis revealed organic compounds that could act as stabilizing agents, suggesting the presence of proteins involved in nanoparticle synthesis. This research is the first to conduct a proteomic analysis of the aqueous filtrate of T. harzianum with applications in the biogenic synthesis of NPs.
