Abstract
BackgroundXylose is the second most abundant carbohydrate in the lignocellulosic biomass hydrolysate. The fermentation of xylose is essential for the bioconversion of lignocelluloses to fuels and chemicals. However the wild-type strains of Saccharomyces cerevisiae are unable to utilize xylose. Many efforts have been made to construct recombinant yeast strains to enhance xylose fermentation over the past few decades. Xylose fermentation remains challenging due to the complexity of lignocellulosic biomass hydrolysate. In this study, a modified genome shuffling method was developed to improve xylose fermentation by S. cerevisiae. Recombinant yeast strains were constructed by recursive DNA shuffling with the recombination of entire genome of P. stipitis with that of S. cerevisiae.ResultsAfter two rounds of genome shuffling and screening, one potential recombinant yeast strain ScF2 was obtained. It was able to utilize high concentration of xylose (100 g/L to 250 g/L xylose) and produced ethanol. The recombinant yeast ScF2 produced ethanol more rapidly than the naturally occurring xylose-fermenting yeast, P. stipitis, with improved ethanol titre and much more enhanced xylose tolerance.ConclusionThe modified genome shuffling method developed in this study was more effective and easier to operate than the traditional protoplast-fusion-based method. Recombinant yeast strain ScF2 obtained in this study was a promising candidate for industrial cellulosic ethanol production. In order to further enhance its xylose fermentation performance, ScF2 needs to be additionally improved by metabolic engineering and directed evolution.
Highlights
Xylose is the second most abundant carbohydrate in the lignocellulosic biomass hydrolysate
While rational metabolic engineering was effective in improving phenotypes of S. cerevisiae strains for xylose fermentation [4], it normally involves the constitutive expression of multiple genes followed by necessary mutagenesis and post-evolutionary engineering
Modified method of genome shuffling Protoplast fusion has been regarded as a traditional and effective way to accelerate strain evolution and been applied in many studies. It suffers from the disadvantages of low efficiency of fusion induced by polyethylene glycol (PEG), labour-intensive and timeconsuming protoplast preparation and fusant regeneration, and fusant instability
Summary
Xylose is the second most abundant carbohydrate in the lignocellulosic biomass hydrolysate. While rational metabolic engineering was effective in improving phenotypes of S. cerevisiae strains for xylose fermentation [4], it normally involves the constitutive expression of multiple genes followed by necessary mutagenesis and post-evolutionary engineering. Considering the complexity of pathway design for rational metabolic engineering, genome shuffling uses recursive genetic recombination analogous to DNA shuffling [6] This strategy was successfully applied in rapid strain improvement of both prokaryotic and eukaryotic cells [7,8]. It is time-saving, easier to operate and has higher gene recombination efficiency
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