Transcription profiling by array Saccharomyces cerevisiae after GGTI-298 treatment.
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ABSTRACT: Global gene expression on yeast cells was determined after a drug induced inhibition of GGTase I by treating yeast cells with GGTI298 (Cabiochem). GGTI298 treated cells were grown for 2 generations to exponential phase in parallel with untreated cells. RNA was extracted, subsequently cDNAs were labelled and hybridized on UHN Y6.4k4 arrays. Replicates with dye-swap of the experiments were generated. Results enabled us to identify the genome-wide effects after drug-induced GGTase I inhibition
Project description:Global gene expression on yeast cells was determined after a drug induced inhibition of FTase by treating yeast cells with FTase Inhibitor I (FTI, Cabiochem). FTI treated cells were grown for 2 generations to exponential phase in parallel with untreated cells. RNA was extracted, subsequently cDNAs were labelled and hybridized on UHN Y6.4k4 arrays. Replicates with dye-swap of the experiments were generated. Results enabled us to identify the genome-wide effects after drug-induced FTase inhibition
Project description:The goal of these experiments was to determine the global expression profile response of budding yeast to a genetic block of Farnesyl Transferase (FTase). We blocked this enzyme using cells that harbour a deletion of the RAM1 gene, which encodes one of the two subunits of FTase. We compared the cells of the strain Delta ram1 with isogenic wild-type cells, grown in parallel cultures. Cells were grown for 2 generations to exponential phase. RNA was extracted, subsequently cDNAs were labelled and hybridized on UHN Y6.4k4 arrays. Replicates with dye-swap of the experiments were generated. Results enabled us to identify the genome-wide effects after genetically-induced FTase block
Project description:We examined the gene expression profiles of CD4+ T-cells isolated from 7 ATL, 12 HAM/TSP and 11 AC (asymptomatic carriers) to identify gene signatures that may be characteristic for these particular diseases. Using gene expression arrays, we identified ~ 1039 immune-related genes that were differentially expressed in CD4+ T cells; using stringent exclusion criteria, a 122 gene signature could be divided into 3 groups: I) ATL-specific; II) common; and III) HAM/TSP-specific markers To better understand the genetic differences between HTLV-1-associated diseases, we examined the gene expression profile of T lymphocytes from patients either suffering from ATL, HAM/TSP or carrying the HTLV-1 virus without any symptoms. Microarray experiments were performed using the human ImmuneArray cDNA array (UHN Microarray Center, University of Toronto). We used the Stratagene Universal Control, labelled Cy5 for all samples in a competitive hybridization.
Project description:Evaluation of the transcriptomic profile of the rabbit embryo along the preimplantation period during in vivo development. Three embryonic stages were used: four cell embryos (H32 post-coïtum); morula (H58 pc) and blastocyst (H90 pc). Keywords: time course rabbit embryo 18 samples
Project description:Comparative transcriptomic analysis: dietary treatment, genetic selection, and muscle fat content We used fish from two divergent lines of rainbow trout, selected for low or high muscle fat content (respectively L and F lines) and fed two different diets, containing either 10% (LE diet) or 23% (HE diet) lipids. For more details about experimental fish and diets, see Kolditz et al., Am J Physiol Regul Integr Comp Physiol 294: R1154âR1164, 2008. There are 4 experimental conditions: L-LE, L-HE, F-LE and F-HE. This experiment contain the data derived from the hybridization of the RNA extracted from individual muscle of these fish. For each membrane, spots with an oligonucleotide signal lower than three times the background level were excluded from the analysis. After this filtering step, signal processing was performed using the vector oligonucleotide data to correct each spot signal according to the actual amount of DNA present in each spot. After correction, the signal was normalized by dividing each gene expression value by the median value of the array. The study consisted of 19 samples total.
Project description:Axis specification in mouse is determined by a sequence of reciprocal interactions between embryonic and extra-embryonic tissues so that a few extra-embryonic genes appear as ‘patterning’ the embryo. Considering these interactions as essential, but lacking in most mammals the genetically driven approaches used in mouse and the corresponding patterning mutants, we examined whether a molecular signature originating from extra-embryonic tissues could relate to the developmental stage of the embryo proper and predict it. To this end, we have profiled bovine extra-embryonic tissues at peri-implantation stages, when gastrulation and early neurulation occur, and analysed the subsequent expression profiles through the use of predictive methods as previously reported for tumour classification. A set of six genes (CALM1, CPA3, CITED1, DLD, HNRNPDL, and TGFB3), half of which had not been previously associated with any extra-embryonic feature, appeared significantly discriminative and mainly dependent on embryonic tissues for its faithful expression. The predictive value of this set of genes for gastrulation and early neurulation stages, as assessed on naive samples, was remarkably high (93%). In silico connected to the bovine orthologues of the mouse patterning genes, this gene set is proposed as a new trait for embryo staging. As such, this will allow saving the bovine embryo proper for molecular or cellular studies. To us, it offers as well new perspectives for developmental phenotyping and modelling of embryonic/extra-embryonic co-differentiation. 20 samples - Amplified material from each embryo (stage 2, stage3, stage4, stage5+) was indirectly labelled using "random" hexamers. One independent target per embryo was generated and hybridised onto one array. 5 measurements per embryonic stage were generated.
Project description:Among the most important regulators of gene expression in bacteria are 'nucleoid-associated proteins'. These proteins alter the topology of the bound DNA by bending, wrapping or bridging it, thus having multiple effects, including transcriptional regulation, on the bacterial cell. Among the best-studied nucleoid proteins are H-NS and Fis, which bind to specific sequences on the DNA. H-NS is a global repressor of gene expression. Fis alters the global conformation of the DNA by introducing branched structures in it; but its effect on gene expression on a genomic scale remains largely unclear.<br><br>Several bacterial transcriptional regulators including H-NS and Fis have been studied using ChIP-chip. However, the higher resolution and dynamic range offered by ChIP-Seq have not been exploited for any bacterial species. By performing ChIP-Seq of these two proteins, we present the first such study in a bacterium. In addition to providing a proof-of-principle for the use of this technology for bacteria, we perform our study at multiple time-points during growth in rich medium, thus generating new insights into how these proteins function under different cellular conditions. Further, by analysing our data in conjunction with newly-generated gene expression and RNA polymerase-chromosome interaction data we provide new interpretation of the genome-scale patterns of the interactions of these proteins to the DNA.
Project description:Here we improved BiTS-ChIP (Bonn et al, Nature Protocols 7, 978-994 (2012)) to identify active enhancer and promoter elements genome wide in the 104 cardiomyocytes that constitute the Drosophila heart tube and represents only ~0.5% of the total cell content of the embryo. A transgenic Drosophila strain expressing nuclear GFP under the control of a cardiac specific enhancer (TinC*>GFP) was used for staged embryo collections at stages 13-14 (10-13h of development). After embryo fixation and dissociation, intact fixed nuclei were fluorescent labelling. Purification of this rare nuclear population was achieved by a two-step sorting procedure, yielding ~98% purity. Chromatin was extracted and used for immunoprecipitation and sequencing (ChIP-seq) to analyze chromatin modifications at promoters (H3K4me3 and H3K27ac) and enhancers (H3K27ac). Two independent biological replicates (from FACS sorting, chromatin preparations and ChIP-Seq) were performed for each mark and sequenced using Illumina HiSeq.
Project description:Nucleosomes are barriers to transcription in vitro, however, their effects on RNA polymerase in vivo are unknown. Here we describe a simple and general strategy to comprehensively map the positions of elongating and arrested RNA Polymerase II (RNAPII) at nucleotide resolution. Our results suggest that nucleosomes present significant, context-specific barriers to RNAPII in vivo that can be tuned by the incorporation of H2A.Z. 8 sets of two replicates each of paired-end and 3 sets of two replicates each of single-end samples were sequenced and analyzed.
Project description:Data presented here show that gene expression in the mouse brain changes during systemic infection relative to a steady state (uninfected control) mouse during systemic Listeria monocytogenes infection. Microarray analysis of gene expression in the brain following i.v. inoculation of 1-2 LD 50 L. monocytogenes showed changes in brain transcriptional activity occurred prior to brain infection. Keywords: infectious diseases model, gene expression profiling for cell signaling/trafficking Mice were injected with 1-2 LD50 wild type L. monocytogenes or with sterile PBS (control) then seven animals from each group were harvested daily for 4 days. From each day, groups of five infected animals were selected for microarray analysis based upon similarities in CFU bacteria in the spleen (day 1), blood (day 2), and brain (days 3 and 4) and were randomly paired with brains harvested from control animals. RNA was extracted from the brain halves using the Atlas⢠Pure Total RNA Labeling System and hybridized to plastic slides with 32P-labeled probes using Atlas⢠Plastic Mouse 5K Microarray (Clontech, Mountain View, CA) according to the manufacturerâs directions. Hybridization to the arrays and phosphorimaging analysis were performed in the OUHSC microarray facility. Phosphorimaging analysis was performed using a Storm⢠optical scanner (Molecular Dynamics Inc., Sunnyvale, CA), and initial background corrections were performed using the Array Vision Software (Imaging Research Inc., St. Catharines, Ontario). Data analyses were performed using Genespring GX v. 7.3 (Agilent Technologies, Palo Alto, CA). Raw signal data were normalized to the median of the entire chip for each sample, and all samples were further normalized per gene to the medians of data from the uninfected samples. Data for each day were analyzed by Mann Whitney Wilcoxen nonparametric test with a p-value cutoff of 0.05. Lists of genes were generated to reflect 2, 3, or 5-fold upregulation and downregulation based on significance. Expression data were incorporated into Ingenuity software (Ingenuity Systems, Inc., Redwood City, CA) to enable further understanding of complex relationships as well as provide further information about key canonical pathways in which the genes of interest functions.