Cost effective processing of wheat straw using solid state fermentation (SSF) would provide a source for value added chemicals from agricultural waste biomass. Fungi natural breakdown lignocellulosic biomass hence could have received a lot of attention. In this study the ability of S. lacrymans to convert straw waste was compared with other Basidiomycetes (Postia placenta, Phanerochaete chrysosporiom, and Schizophyllum commune). S. lacrymans out performed the other Basidiomycetes both in its growth (as measured by ergosterol and fatty acid production (linoleic acid);18:2n6c) , and in the comounds released which included; total soluble phenols, total reducing sugars, and low molecular organic chemicals (MW<400). Non-enzymatic breakdown requiring the presence of Fe2+ was also demonstrtated and influenced by the production of quinone and low molecular organic acid. The amount of the fungal extract used and the concentration of chelator/reducing agents also affected the production of Fe2+. Changes in the lignocellulose structure was also detected as key functional group, such as the pyranose ring and aromatic skeletal vibration were significantly reduced following culture with S. lacrymans and a significant reduction in mass was measured. Iron reductase genes IR1 and IR2 suspected to be involved in the lignocellulose degradation were cloned. It seems that these genes are more closely related to the cellulose binding module (CBM) family instead of cellobiose dehydrogenase (CDH) genes as first suspected. IR1 has an open reading frame of 774 bp which encoding 258 amino acid (55 kDa), whilst IR2 642 bp encoding 214 amino acid ( 49 kDa). The IR1 gene contains a CBM1 domain which is lacking in IR2. Gene expression analysis using qRT-PCR showed that in the early stage of fungal growth, the level of IR2 genes expression was higher than IR1 while IR1 became more dominant in the latter stages of culture. The time at which these genes are highly expressed correlated with the release of soluble and aromatic phenolic compounds. The functionality of the recombinant IR1 and IR2 on the decomposition of lignocellulose was shown using several assays including; iron reductases, nitrated lignin and the reduction of electron acceptor (DCPIP). In addition, using both synthetic and nature sources of cellulose or lignocellulose (Avicel and wheat straw powder) the recombinant IR proteins were shown to break down cellulose. This suggested that these enzymes represent a significant addition to those currently used within biomass based biorefineries.

The rapid depletion of fossil fuel reserves and concurrent increase in global temperatures has resulted in global demand for the production of alternative environmentally friendly fuels. First-generation biofuels that utilise cash crops for the extraction of fermentable sugars currently exist, but are highly controversial due to socioeconomic and environmental reasons such as diverting food production or deforestation. Therefore, second-generation biofuels that utilise lignocellulosic waste materials are a more attractive prospect. In Europe, lignocellulosic biomass wastes such as wheat straw, display great potential for the production of alternative energy sources such as bioethanol for transportation. Conversion to this biofuel requires microorganisms that will effectively utilise the constituent sugars to produce a high yield of product. Saccharomyces cerevisiae (S. cerevisiae) strains possess the most desirable phenotypes for this objective. However, the components of wheat straw are difficult to break down, therefore pretreatment is required. Pretreatment methods vary but often utilise various chemicals that produce compounds that are inhibitory to yeast. This affects the efficiency of fermentations. The focus of this work is on formic acid and a synthetic media containing the main inhibitor compounds released during pre-treatment of steam exploded wheat straw. Six pair-wise F1 crosses between four distinct parental S. cerevisiae clean lineage populations have been generated previously by Cubillos et al., 2009. The 96 F1 progeny from each cross have been assayed for tolerance phenotypes in order to determine QTLs (Quantitative Trait Loci), which will enable us to map genes contributing to the multi-genic trait of inhibitor tolerance. Overall, three QTLs were identified for formic acid and five QTLs were identified from the synthetic inhibitor mix. Candidate genes were selected from the QTL analysis and were tested by performing reciprocal hemizygosity assays to determine which genes are responsible for inhibitor resistance to enable the development of yeast strains suitable for second-generation biofuel production.