The production of biofuels from plant biomass is dependent on the availability of enzymes that can hydrolyze the plant cell wall polysaccharides to their monosaccharides. These enzyme mixtures are formed by microorganisms but their native compositions and properties are often not ideal for application. Genetic engineering of these microorganisms is therefore necessary, in which introduction of DNA is an essential precondition. The filamentous fungus Trichoderma reesei-the main producer of plant-cell-wall-degrading enzymes for biofuels and other industries-has been subjected to intensive genetic engineering toward this goal and has become one of the iconic examples of the successful genetic improvement of fungi. However, the genetic manipulation of other enzyme-producing Trichoderma species is frequently less efficient and, therefore, rarely managed. In this chapter, we therefore describe the two potent methods of Trichoderma transformation mediated by either (a) polyethylene glycol (PEG) or (b) Agrobacterium. The methods are optimized for T. reesei but can also be applied for such transformation-resilient species as T. harzianum and T. guizhouense, which are putative upcoming alternatives for T. reesei in this field. The protocols are simple, do not require extensive training or special equipment, and can be further adjusted for T. reesei mutants with particular properties.
Keywords: Agrobacterium tumefaciens; Cellulolytic fungus; Plant cell wall degrading enzymes; Protoplasts; RUT-C30; Rhizobium radiobacter; Trichoderma guizhouense; Trichoderma harzianum; Trichoderma reesei; Vector.