DNA replication profiling by BrdU Immunoprecipitation of S.cerevisiae wild-type versus H3K37R mutant under over-expression of MCM activators.
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina HiSeq 4000
ORGANISM(S): Saccharomyces cerevisiae
SUBMITTER: Namshik Han
PROVIDER: E-MTAB-9999 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Wild-type and isogenic H3K37R yeast cultures were sinchronized in G1 with alpha factor. Cell were released into the cycle in the presence of 200mM Hydroxyurea and 0.5mg/ml BrdU. After 1h 15 minutes at 30C, replication was stopped by addition of NaN3 and cells were processed for DNA immunoprecipitation with anti BrdU antibody.
2021-04-14 | E-MTAB-9097 | biostudies-arrayexpress
Project description:Cells were grown to saturation in YPD (YEP + 2% glucose) for 24 hours, diluted into YPA (YEP + 2% potassium acetate) at OD600= 0.3 and grown over night at 30C. Cells were washed with sterilized water the next day and re-suspended in SPII medium (0.3% potassium acetate, pH = 7.0) at OD600= 1.9 to induce sporulation. Cells were sporulated at room temperature or 30C as indicated. Sporulation medium containing benomyl was always prepared freshly on the day of the experiment following the directions in {Shonn, 2000 #90}. Briefly, DMSO (dimethyl sulfoxide, Sigma-Aldrich) or benomyl [Methyl 1-(butylcarbamoyl)-2-benzimidazolecarbamate, Sigma-Aldrich; 30 mg/ml stock in DMSO] was dissolved in near-boiling SPII medium to avoid precipitation. The medium was then allowed to slowly cool to 30C or room temperature. At the time of drug treatment, cells were filtered and immediately re-suspended in the medium containing benomyl or DMSO. Keywords = benomyl sporulation saccharomyces microtubule spindle checkpoint Keywords: ordered
2004-10-18 | GSE1693 | GEO
Project description:To investigate the impact of Rap1 binding on nearby gene expression during the transition from vegetative growth to meiosis, we measured RNA abundance in cells depleted (plus doxycycline) or not depleted (minus doxycycline) of Rap1. RNA abundance was measured in depleted and non-depleted condition in respiratory cells (pre-meiotic/YPA 20 hours) and in cells undergoing synchronous meiosis/sporulation in the budding yeast Saccharomyces cerevisiae. There is growing recognition that the binding of a transcription factor near a gene does not always indicate regulatory function, and further that a single factor may function to either activate or repress its targets depending on the cellular context. We examined these issues through a series of experiments involving the S. cerevisiae transcription factor Rap1, and its function throughout critical metabolic and developmental transitions between vegetative growth, respiratory growth, meiosis and sporulation. We simultaneously monitored the expression of all genes and the genomic binding locations of Rap1 throughout the timecourse. Genes downstream of Rap1 binding were activated and repressed dynamically, but a change - or lack of change - in Rap1 binding status was not predictive of activation, repression, or no change in regulation. Despite this, we show that Rap1 is required, at a given point in time, for both activation and repression of different gene targets, within the same cell. Specification of the transcriptional consequences of Rap1 binding is thus highly promoter-specific. The presence of other transcription factor binding motifs, the subtype of Rap1 motif, and the underlying chromatin structure of the promoter cannot fully account for the observed transcriptional outcomes. There is a better accounting for the dynamic binding behavior of Rap1 including specification of an expanded meiotic target set through a Tup1- dependent nucleosome-loss mechanism. The variable and dynamic association between binding and transcription in this simple unicellular system portends a similarly volatile relationship in more complex eukaryotes. Biological interpretations of transcription factor occupancy should be made cautiously and in conjunction with supporting data obtained under the precise condition of interest. SK1 yeast strain SHy20 (MATa/MATalpha lys2/lys2 ura3/ura3 ho::LYS2/ho::LYS2 leu2::hisG/leu2::hisG his4x/his4b URA3::CMV-tetTA/URA3::CMV-tetTA RAP1promoter::kanR-tetO7-TATA/RAP1promoter::kanR-tetO7-TATA). An overnight culture of YPD was used to innoculate a YPA culture (minus doxycycline; Rap1 on). The YPA culture was grown for 8 hours and then split into minus doxycyline (Rap1 on) and plus doxycylcine (Rap1 depleted) samples. Minus and plus doxycyline samples were collected after an additional 12 hours in YPA (respiratory growth/pre-meiotic; YPA 20 hours) and 0 hr, 3hr, 6hr, 9hr and 12hr after transfer to sporulation media (SM) with or without doxycycline. 3 separate biological replicates were used for each condition/time-point. Labeled total RNA from Rap1 depeleted and non-depleted samples were competitively hybridized to yeast whole genome PCR based spotted arrays (resolution ~1kb). The Rap1-depletion ChIP and RNA abundance/expression analysis were carried out on the same biological samples. The three replicates of Rap1-depletion RNA abundance/expression analysis correspond to replicates 2, 3, and 5 of the Rap1-depletion Rap1 ChIP experiment.
2016-09-01 | E-GEOD-75259 | biostudies-arrayexpress
Project description:Cells were grown to saturation in YPD (YEP + 2% glucose) for 24 hours, diluted into YPA (YEP + 2% potassium acetate) at OD600= 0.3 and grown over night at 30C. Cells were washed with sterilized water the next day and re-suspended in SPII medium (0.3% potassium acetate, pH = 7.0) at OD600= 1.9 to induce sporulation. Cells were sporulated at room temperature or 30C as indicated. Sporulation medium containing benomyl was always prepared freshly on the day of the experiment following the directions in {Shonn, 2000 #90}. Briefly, DMSO (dimethyl sulfoxide, Sigma-Aldrich) or benomyl [Methyl 1-(butylcarbamoyl)-2-benzimidazolecarbamate, Sigma-Aldrich; 30 mg/ml stock in DMSO] was dissolved in near-boiling SPII medium to avoid precipitation. The medium was then allowed to slowly cool to 30C or room temperature. At the time of drug treatment, cells were filtered and immediately re-suspended in the medium containing benomyl or DMSO.
2008-06-11 | E-GEOD-1693 | biostudies-arrayexpress
Project description:Experimental factor: Time after the temperature shift from 30C to 37C or 30C to 48C. Conidia (5x106 /ml) from Aspergillus fumigatus Af293 were incubated in the CM medium for germination (~ 17 hrs) at 30C. The culture was transferred to a water bath of 37C or 48C for continued growth. Samples were taken after the defined time (i.e. 0, 15, 30, 60, 120, and 180 min.).
2005-05-15 | E-MEXP-332 | biostudies-arrayexpress
Project description:To investigate transcriptome dynamics during the transition from vegetative growth to meiosis, we measured RNA abundance in vegetative cells (YPD), respiratory cells (pre-meiotic/YPA 8 hours), and in cells undergoing synchronous meiosis/sporulation in the budding yeast Saccharomyces cerevisiae. There is growing recognition that the binding of a transcription factor near a gene does not always indicate regulatory function, and further that a single factor may function to either activate or repress its targets depending on the cellular context. We examined these issues through a series of experiments involving the S. cerevisiae transcription factor Rap1, and its function throughout critical metabolic and developmental transitions between vegetative growth, respiratory growth, meiosis and sporulation. We simultaneously monitored the expression of all genes and the genomic binding locations of Rap1 throughout the timecourse. Genes downstream of Rap1 binding were activated and repressed dynamically, but a change - or lack of change - in Rap1 binding status was not predictive of activation, repression, or no change in regulation. Despite this, we show that Rap1 is required, at a given point in time, for both activation and repression of different gene targets, within the same cell. Specification of the transcriptional consequences of Rap1 binding is thus highly promoter-specific. The presence of other transcription factor binding motifs, the subtype of Rap1 motif, and the underlying chromatin structure of the promoter cannot fully account for the observed transcriptional outcomes. There is a better accounting for the dynamic binding behavior of Rap1 including specification of an expanded meiotic target set through a Tup1- dependent nucleosome-loss mechanism. The variable and dynamic association between binding and transcription in this simple unicellular system portends a similarly volatile relationship in more complex eukaryotes. Biological interpretations of transcription factor occupancy should be made cautiously and in conjunction with supporting data obtained under the precise condition of interest. SK1 yeast strain SHy002 (MATa/MATα ho::LYS2/ho::LYS2 leu2::hisG/leu2::hisG lys2/lys2 ura3/ura3). Meiosis timecourse with samples collected during vegetative growth (YPD), respiratory growth (pre-meiotic/YPA 8 hours) and 0 hr, 1.5hr, 3hr, 4.5hr, 6hr, 9hr and 12hr after transfer to sporulation media (SM). 7 separate biological replicates were used for each condition/time-point. Labeled total RNA from each sample and a labeled genomic DNA common reference were competitively hybridized to yeast whole genome PCR based spotted arrays (resolution ~1kb). The Rap1 ChIP, RNA abundance/expression analysis, and FAIRE analysis were all carried out on the same biological samples.
2016-09-01 | E-GEOD-75257 | biostudies-arrayexpress
Project description:To investigate the genome-wide binding dynamics of Rap1 during the transition from vegetative growth to meiosis, we performed Rap1 ChIP-chip experiments in vegetative cells (YPD), respiratory cells (pre-meiotic/YPA 8 hours), and in cells undergoing synchronous meiosis/sporulation in the budding yeast Saccharomyces cerevisiae. There is growing recognition that the binding of a transcription factor near a gene does not always indicate regulatory function, and further that a single factor may function to either activate or repress its targets depending on the cellular context. We examined these issues through a series of experiments involving the S. cerevisiae transcription factor Rap1, and its function throughout critical metabolic and developmental transitions between vegetative growth, respiratory growth, meiosis and sporulation. We simultaneously monitored the expression of all genes and the genomic binding locations of Rap1 throughout the timecourse. Genes downstream of Rap1 binding were activated and repressed dynamically, but a change - or lack of change - in Rap1 binding status was not predictive of activation, repression, or no change in regulation. Despite this, we show that Rap1 is required, at a given point in time, for both activation and repression of different gene targets, within the same cell. Specification of the transcriptional consequences of Rap1 binding is thus highly promoter-specific. The presence of other transcription factor binding motifs, the subtype of Rap1 motif, and the underlying chromatin structure of the promoter cannot fully account for the observed transcriptional outcomes. There is a better accounting for the dynamic binding behavior of Rap1 including specification of an expanded meiotic target set through a Tup1- dependent nucleosome-loss mechanism. The variable and dynamic association between binding and transcription in this simple unicellular system portends a similarly volatile relationship in more complex eukaryotes. Biological interpretations of transcription factor occupancy should be made cautiously and in conjunction with supporting data obtained under the precise condition of interest. SK1 yeast strain SHy002 (MATa/MATα ho::LYS2/ho::LYS2 leu2::hisG/leu2::hisG lys2/lys2 ura3/ura3). Meiosis timecourse with samples collected during vegetative growth (YPD), respiratory growth (pre-meiotic/YPA 8 hours) and 0 hr, 1.5hr, 3hr, 4.5hr, 6hr, 9hr and 12hr after transfer to sporulation media (SM). 7 separate biological replicates were used for each condition/time-point. Labeled ChIP and input samples were competitively hybridized to yeast whole genome PCR based spotted arrays (resolution ~1kb). The Rap1 ChIP, RNA abundance/expression analysis, and FAIRE analysis were all carried out on the same biological samples. The two Mock Rap1 ChIP timecourse replicates correspond to timecourse replicates 3 and 4.
2016-09-01 | E-GEOD-75261 | biostudies-arrayexpress
Project description:To identify meiotic non-coding RNAs, a DNA strand-specific transcript profiling experiment using cultures of Saccharomyces cerevisiae in YPD, YPA and SPII medium and tiling microarrays (GeneChips) covering both strands of the strain S288c complete genome was carried out.
2010-02-01 | E-TABM-915 | biostudies-arrayexpress
Project description:JN54 (wild-type) cells were incubated in YPD medium at 30 degrees C to a logarithmic phase (OD660=1), followed by treatment with mild heat-shock at 43 degrees C for 30 min in pre-warmed (43 degrees C) 100ml of YPD medium using 500 ml Erlenmeyer flask. JN54 (wild-type) cells were incubated in YPD medium at 30 degrees C to a logarithmic phase (OD660=1), followed by treatment with mild heat-shock at 43 degrees C for 60 min in pre-warmed (43 degrees C) 100ml of YPD medium using 500 ml Erlenmeyer flask. Keywords: Stress response
2005-09-21 | GSE3316 | GEO
Project description:To investigate the dynamics of the meiotic transcriptome and to ensure meiotic synchrony of our samples, we measured RNA abundance in vegetative cells, respiratory (pre-meiotic) cells and in synchronously sporulating cells at various times after transfer to sporulation media (SM). SK1 yeast strain SHy002 (MATa/MATα ho::LYS2/ho::LYS2 leu2::hisG/leu2::hisG lys2/lys2 ura3/ura3)<br> Meiotic Time Course with samples collected during vegetative growth (YPD), respiratory growth (pre-meiotic/YPA 8 hours) and 0 hr, 1.5hr, 3hr, 4.5hr, 6hr, 9hr and 12hr after transfer to sporulation media (SM).<br> 3 separate biological replicates were used for each condition/time-point.<br> Total RNA from each sample was hybridized competitively with a genomic DNA common reference.<br> The expression analysis was carried out from the same biological samples as the FAIRE analysis.<br>
2010-05-22 | E-GEOD-18181 | biostudies-arrayexpress