BSA by Sequencing
Bulk Segregant Analysis by High Throughput Sequencing Reveals a Novel Xylose Utilization Gene from Saccharomyces cerevisiae
Fermentation of xylose is of fundamental importance for the efficient production of ethanol from lignocellulosic biomass sources.  Native Saccharomyces cerevisiae strains have traditionally been thought incapable of either fermentative or non-fermentative growth on xylose despite their high efficiency ethanol production from fermentation of hexoses, although population genetic approaches have found a few non-engineered strains that show a modest xylose utilization phenotype.  Genetically engineered xylose-fermenting strains of S. cerevisiae have been constructed, yet no strain, either natural or engineered, has been identified that can ferment xylose as efficiently as glucose.  Here, we used a medium throughput screen to identify Saccharomyces strains that can increase in optical density when xylose is present as the sole carbon source; we identified 38 strains that have this xylose utilization phenotype including S. cerevisiae, other sensu stricto members, and hybrids between them.  All of the S. cerevisiae positive strains we identified are wine yeasts, and of those that we were able to generate viable spores from, the xylose phenotype segregates as a single gene trait.  We mapped this gene by Bulk Segregant Analysis (BSA) using tiling microarrays and high-throughput sequencing.  The gene is a putative xylitol dehydrogenase, which we named XDH1, and is located in the subtelomeric region of the right end of chromosome XV in a region not present in the S288c reference genome.  We further characterized the xylose phenotype by performing gene expression microarrays and by genetically dissecting the endogenous Saccharomyces xylose pathway. We have demonstrated that natural Saccharomyces cerevisiae yeasts are capable of utilizing xylose as the sole carbon source, characterized the genetic basis for this trait as well as the endogenous xylose utilization pathway, and provided Bulk Segregant Analysis as a novel application of high throughput sequencing.