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Xylose utilization timecourse

ABSTRACT: We have identified a novel xylose-utilization phenotype controlled by a single gene, and we sought to characterize how both the presence and absence of xylose in the presence or absence of the trait affected the S. cerevisiae transcriptional program over time. To do so, we measured global steady state mRNA levels on three pairs of sister spores from a Simi White strain that was backcrossed twice to S288c. Each pair of spores was from an independent tetrad, and contained one xylose utilizing ("positive", GSY2465, 2466, 2469) and one xylose non-utilizing spore ("negative", GSY2464, 2467, 2470). We grew each of the six spores in the presence or absence of xylose for 72 hours with samples taken every 8 hours. We then assayed relative RNA abundance versus a pooled reference, containing equimolar amounts of each sample, using Agilent yeast catalog arrays.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE19116 | GEO | 2010/04/09

SECONDARY ACCESSION(S): PRJNA123739

REPOSITORIES: GEO

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Xylose utilization timecourse

Project description:We have identified a novel xylose-utilization phenotype controlled by a single gene, and we sought to characterize how both the presence and absence of xylose in the presence or absence of the trait affected the S. cerevisiae transcriptional program over time. To do so, we measured global steady state mRNA levels on three pairs of sister spores from a Simi White strain that was backcrossed twice to S288c. Each pair of spores was from an independent tetrad, and contained one xylose utilizing ("positive", GSY2465, 2466, 2469) and one xylose non-utilizing spore ("negative", GSY2464, 2467, 2470). We grew each of the six spores in the presence or absence of xylose for 72 hours with samples taken every 8 hours. We then assayed relative RNA abundance versus a pooled reference, containing equimolar amounts of each sample, using Agilent yeast catalog arrays. time_series_design

2010-04-09 | E-GEOD-19116 | biostudies-arrayexpress

Bulk Segregant Analysis Reveals a Novel Xylose Utilization Gene from Saccharomyces cerevisiae

Project description:To determine the genomic location of a gene that permits xylose utilization we conducted bulk segregant analysis (BSA) using Affymetrix yeast tiling arrays. BSA works by taking advantage of DNA sequence polymorphisms between different strains and the fact that it is relatively easy to pool large numbers of meiotic spore products (segregants) in yeast. Pooling segregants based on their phenotype allows the region of the genome responsible for the phenotype to be detected. This is because DNA polymorphisms in regions unlinked to the locus causing the phenotype will segregate randomly and be “evened” out, while around the genomic region of interest, sequences or polymorphisms responsible for the trait will be present in all positive segregants, and absent in all negative segregants. In our case, a Simi White wine strain (S. cerevisiae) carrying the locus responsible for xylose utilization was crossed to a laboratory strain of Saccharomyces cerevisiae; this strain was estimated to carry DNA polymorphisms relative to the laboratory strain at a level of approximately .5%. Spores from the Simi White / S288c diploid were screened for the xylose utilization phenotype and 39 positive spores were combined into one pool and 39 negative spores into another pool, and genomic DNA (gDNA) was isolated from each pool. We then hybridized the positive and negative gDNA pools to tiling microarrays that were based on the S288c reference genome with the expectation that regions of the genome derived from Simi White will hybridize less robustly to the array because of the DNA polymorphisms between Simi White and S288c. Log2 ratios of probe intensities were calculated (negative/positive), and a peak appeared in the chromosome XV right subtelomeric region that corresponds to less robust hybridization to the microarray of the positive pool gDNA coming from this region of the genome

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Project description:Array hybridization data for 611 yeast segregants to determine genotype at each marker. DNA from each segregant was digested with DNAseI, labeled with Cy5-ddATP, and hybridized individually to a custom agilent 8x15k, genotyping array . Note: Tetrad refers to the tetrad from the cross of YJM145 x S288c that each segregant was dissected from. Tag ID refers to unique yeast barcode has been integrated at the ho locus in a segregant.

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Project description:HMF and furfural were pulse added to xylose-utilizing Saccharomyces cerevisiae during either the glucose consumption phase or the xylose consumption phase. Transcriptome samples were collected before and one hour after pulsing of inhibitors. Three biological replicates from each conditions analyzed.

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