Challenges of strain development and clone selection for bioethanol production from lignocellulose
Conference poster, 2013
Lignocellulosic biomass is one of the most promising materials for bioethanol production because it does not compete with food crops and it is widely distributed around the world. However, when using lignocellulosic biomass as raw material, inhibitory compounds derived from cellulose, hemicellulose and lignin degradation during pretreatment are also found in the fermentation media. It is well known that the most commonly used microorganism for ethanol production is Saccharomyces cerevisiae. Wild type S. cerevisiae, however, is not able to ferment xylose, which could constitute up to 40% of the lignocellulose and therefore recombinant xylose utilizing strains have been developed. Yeasts strains to be used for second-generation bioethanol production have to cope with challenging conditions that are inherent to the industrial process such as high concentration of inhibitory products, simultaneous use of different carbon sources and growth conditions that are not well controlled. Tolerance to these multiple stresses is likely to be a complex phenotype involving several cellular mechanisms and it could be difficult to perform efficient metabolic engineering. In this context, one of the most promising strategies for developing industrial strains is evolutionary engineering that includes evolution and recombination introducing genetic variability over many generations.
Evolved S. cerevisiae strains engineered for xylose fermentation employed in this study have been subjected to targeted engineering for introducing a barcode in order to be able to verify their origin, which also provokes random events in the population of cells. Screening after evolution or targeted engineering is challenging because of the high variability introduced during those events. Selection has to be performed carefully in order to select the best clones with best properties for a specific purpose. Furthermore, difficulties in applying novel technics such as next generation sequencing or multiomic analysis in industrial strains result from their genetic complexity such as polyploidy.
In this work, mixed populations obtained by evolutionary engineering and different clones obtained after barcoding (Figure 1) are tested and evaluated in ethanol production processes from lignocellulosic hydrolysates. Differences between clones regarding xylose fermentation capability and ability to ferment in lignocellulosic hydrolysates are elucidated.