Project description:We developed an artificial genome evolution system, which we termed ‘TAQing’, by introducing multiple genomic DNA double-strand breaks using a heat-activatable endonuclease in mitotic yeast. The heat-activated endonuclease, TaqI, induced random DSBs, which resulted in diverse types of chromosomal rearrangements including translocations. Array comparative genomic hybridization (aCGH) analysis was performed with cell-fused Saccharomyces cerevisiae strains induced genome evolution by TAQing system. Some of copy number variations (CNVs) induced by massive genome rearrangements were detected in the TAQed yeast strains.
Project description:Single-stranded DNA breaks (nicks) were tagged by biotin-dCTP in wild-type budding yeast cells and mapped by nick-ChIP coupled to whole-genomic microarrays.
Project description:Telomere chromatin structure is pivotal for maintaining genome stability by regulating the binding of telomere-associated proteins and inhibition of a DNA damage response. In yeast, the silent information regulator (Sir) proteins bind to terminal telomeric repeats and to subtelomeric X-elements resulting in histone deacetylation and transcriptional silencing. Herein, we show that sir2 mutant strains display a very specific loss of a nucleosome residing in the X-element. Most yeast telomeres contain an X-element and the nucleosome occupancy defect in sir2 mutants is remarkably consistent between different telomeres.
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