Project description:We used ChIP-seq to determine the whole-genome enrichment of histone H3 threonine 11 phosphorylation (H3 T11ph) during Saccharomyces cerevisiae meiosis. S. cerevisiae SK1 cells were synchronized for meiotic entry and 3 and 4 hour meiotic samples were obtained. As H3 T11ph is dependent on the formation of meiotic double strand breaks (DSBs), a negative control ChIP-seq sample was obtained from a strain lacking DSBs (spo11-yf). Concurrently, ChIP-seq was carried out for histone H3 as a control for comparision.
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:SPO11-promoted DNA double-strand breaks (DSBs) formation is a crucial step for meiotic recombination, and it is indispensable to detect the broken DNA ends accurately for dissecting the molecular mechanisms behind. Here, we report a novel technique, named DEtail-seq (DNA End tailing followed by sequencing), that can directly and quantitatively capture the meiotic DSB 3’ overhang hotspots at single-nucleotide resolution.
Project description:We are investigating the transcriptional response of yeast to treatment with enediynes or gamma radiation, which generate different extents of double or single strand breaks in DNA. We used microarrays to detail the global programme of gene expression underlying the DNA damage response in yeast Experiment Overall Design: Yeast were grown to mid log phase and treated with enediynes or gamma radiation (in biological triplicate) resulting in similar extents of cell killing. The responses were compared to each other and we have deciphered a gene expression profile that is specific for double and single strand breaks in DNA.
Project description:DNA topoisomerases are required to resolve DNA topological stress. Despite this essential role, abortive topoisomerase activity generates aberrant protein-linked DNA breaks, jeopardising genome stability. Here, to understand the genomic distribution and mechanisms underpinning topoisomerase-induced DNA breaks, we map Top2 DNA cleavage with strand-specific nucleotide resolution across the S. cerevisiae and human genomes—and use the meiotic Spo11 protein to validate the broad applicability of this method to explore the role of diverse topoisomerase family members. Our data characterises Mre11-dependent repair in yeast and defines two strikingly different fractions of Top2 activity in humans: tightly localised CTCF-proximal, and broadly distributed transcription-proximal, the latter correlated with gene length and expression. Moreover, single nucleotide accuracy reveals the influence primary DNA sequence has upon Top2 cleavage—distinguishing sites likely to form canonical DNA double-strand breaks (DSBs) from those predisposed to form strand-biased DNA single-strand breaks (SSBs) induced by etoposide (VP16) in vivo. This data set contains maps of Top2 CCs in the S. cerevisiae genome, generated by CC-seq of BY4741 cells -/+ etoposide (VP16).
