RNA-seq of ChI-D2RKO mouse nucleus accumbens following a progressive ratio schedule of reinforcement of intravenous cocaine or saline
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Bioanalyzer, Nanodrop,, Illumina NovaSeq 6000, UC Irvine HPC
ORGANISM(S): Mus musculus
SUBMITTER: Robert Lewis
PROVIDER: E-MTAB-14303 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Mice were placed in a novel home cage and habituated for 2 hours. They were then injected with saline or cocaine (20 mg/kg ip). After 1 hour, mice were rapidly sacrificed via cervical dislocation, their brains were promptly dissected, and were rapidly frozen in 2-methylbutane. Tissue punches from the dorsomedial striatum were taken and RNA was isolated using Trizol following the manufactures recommendations.
2020-04-22 | E-MTAB-8589 | biostudies-arrayexpress
Project description:We analyzed the RNA expression profiles of Parkinson's model (6-hydroxydopamine lesion, 6-OHDA) mice in two different genotypes, WT and iMSN-D2RKO mice. Mutant iMSN-D2RKO mice were generated by mating D2Rflox/flox males with D2Rflox/flox females carrying the CRE under the control of the D1 receptor promoter (D1)-Cre. This way, we generated the D2Rflox/flox/D1-Cre/+ line; the D2Rflox/flox mice are thereafter called WT in our study. The D1R gene is expressed during development in MSNs precursors allowing the deletion of D2R as previously shown selectively in iMSNs. The RNA expression profiles were analyzed in three experimental groups: 1) sham-lesioned (sham); 2) 6-OHDA-lesioned (6-OHDA); and 3) 6-OHDA lesioned/L-DOPA-treated (15 mg/kg ). (L-DOPA). Mice of both genotypes were sacrificed 1h after the treatment and brains were flash frozen. Punches of the dorsolateralstriatum(DLS) were obtained on cryostat sections and processed for RNA extraction to prepare the library.
2023-06-06 | E-MTAB-11848 | biostudies-arrayexpress
Project description:Perfluorooctanoic acid (PFOA) is one of the most used perfluorinated compounds in numerous applications and can be detected in environmental samples from around the globe. The aquatic environment is an important site for PFOA deposit. Nevertheless, the exact mode of action and its resulting toxicological effects on aquatic organisms remain largely unknown. To gain a more extensive understanding of the mode of action of teleost PFOA toxicity, transcriptomics, proteomics, biochemical parameters and reproduction were integrated in the present study. Male and female zebrafish were exposed to nominal concentrations of 0.1; 0.5 and 1 mg/l PFOA for 4 and 28 days resulting in an accumulation which was higher in males compared to females. These gender-related differences were likely caused by different elimination rates due to distinct hormone levels and differences in transport activity by solute carriers. The general mode of action of PFOA was believed to be an increase of the mitochondrial membrane permeability which caused effects on the electron transport system at the biochemical level and resulted in alterations of the oxidative phosphorylation, oxidative stress and apoptosis at the gene transcript and protein level. As a consequence, evidence for the replacement of the affected cells and organelles to sustain tissue homeostasis was found at the molecular level. The higher energy demand, due to these adverse effects, was provided by lowering the glycogen stores. Despite this increase in metabolic expenditure, no effects on reproduction were found indicating that the fish seemed to cope with exposure to the tested concentrations of PFOA. Adult zebrafish (Danio rerio) were exposed to nominal concentrations of 0mg/l; 0.1mg/l; 1mg/l PFOA (perfluorooctanoic acid) for 28 days. Three different 25 litre aquaria per exposure concentration were used resulting in 3 biological replicates with each aquarium containing 8 male and 8 female zebrafish. The livers of 6 male fish and 6 female fish were pooled separately and snap frozen in liquid nitrogen. A reference sample was made by pooling equal amounts of RNA from all samples. A carriage wheel design was used in which all samples were connected to the reference sample and the main contrasts of interest were made directly on the same microarrays as frequently as possible. This design resulted in technical triplicates of each sample.
2013-07-20 | E-GEOD-19908 | biostudies-arrayexpress
Project description:Changes in gene expression contribute to the long-lasting regulation of the brain's reward circuitry seen in drug addiction, however, the specific genes regulated and the transcriptional mechanisms underlying such regulation remain poorly understood. Here, we used chromatin immunoprecipitation coupled with promoter microarray analysis to characterize genome-wide epigenetic changes in the mouse nucleus accumbens, a crucial brain reward region, after repeated cocaine administration. Our findings reveal several interesting principles of gene regulation by cocaine and of the role of Î?FosB and CREB, two prominent cocaine-induced transcription factors, in this brain region. Mice were treated with cocaine or saline. Chromatin immunoprecipitation was performed for acetylated histone H3 and H4 as described previously (Kumar et al., 2005) with minor modifications. Immunoprecipitated DNA was amplified via ligation-mediated PCR and hybridized to Nimblgen mouse MM5 promoter arrays. 2 biological replicates per condition.
2010-05-18 | E-GEOD-16183 | biostudies-arrayexpress
Project description:Changes in gene expression contribute to the long-lasting regulation of the brain's reward circuitry seen in drug addiction, however, the specific genes regulated and the transcriptional mechanisms underlying such regulation remain poorly understood. Here, we used chromatin immunoprecipitation coupled with promoter microarray analysis to characterize genome-wide epigenetic changes in the mouse nucleus accumbens, a crucial brain reward region, after repeated cocaine administration. Our findings reveal several interesting principles of gene regulation by cocaine and of the role of Î?FosB and CREB, two prominent cocaine-induced transcription factors, in this brain region. Mice were treated with cocaine or saline. Chromatin immunoprecipitation was performed for methylated histone H3 lysine 9 and 27, â??FosB, and phospho-CREB as described previously (Kumar et al., 2005) with minor modifications. Immunoprecipitated DNA was amplified via ligation-mediated PCR and hybridized to Nimblgen mouse MM8 promoter arrays. 2-3 biological replicates per condition.
2010-05-18 | E-GEOD-16184 | biostudies-arrayexpress
Project description:Several recent reports show that the cerebral cortex in humans and animals with altered expressions of Wnt/cadherin network-associate molecules display cytoarchitectural abnormalities reminiscent of cortical dysplasias seen in some (mouse-, rat-, and monkey-based) animal models of prenatal cocaine exposure. Therefore, we employed oligo microarrays followed by real-time RT-PCR to compare expressions of genes involved in Wnt and cadherin systems in the cerebral wall of 18-day-old (E18) fetuses from cocaine-treated (20 mg/kg cocaine, s.c., b.i.d., E8-18) and drug-naive (saline, s.c.) mice. The pregnant mice chronically treated with cocaine in the above-described manner represent one of the animal models producing offspring with widespread cortical dysplasias. Out of more than 150 relevant genes in the arrays, 32 were upregulated and 9 were downregulated in cocaine-exposed fetuses. The majority of these genes (30 out of 41) were similarly affected in the frontal and occipital regions of the cerebral wall. We also used Western immunoblotting to examine the ability of cocaine to regulate the protein levels of beta-catenin, the key functional component of both Wnt and cadherin systems. While the total cell levels of beta-catenin were increased throughout the cerebral wall of cocaine-exposed fetuses, its nuclear (gene-transcription driving) levels remained unaltered. This suggests a transcription-unrelated role for cocaine-induced upregulation of this protein. Overall, our findings point to an intriguing possibility that that cerebral cortical dysplasias observed in several animal models of prenatal cocaine exposure may be at least in part related to alterations in the Wnt/cadherin molecular network. It has been demonstrated that exposure to cocaine increases cell death in the fetal CNS. To examine the molecular mechanisms of this effect, we employed mouse oligo microarrays followed by real-time reverse transcriptase-polymerase chain reaction (real-time RT-PCR) to compare expressions of apoptosis-related genes in the cerebral wall of 18-day-old (E18) fetuses from cocaine-treated (20 mg/kg cocaine, s.c., b.i.d., E8th-E18th) and drug-naive (saline, s.c.) mice. Out of approximately 400 relevant genes in the arrays, 53 showed alterations in expression in cocaine-exposed fetuses. Upregulation was observed in 35 proapoptotic and 8 antiapoptotic genes; 4 proapoptotic and 6 antiapoptotic genes were down-regulated. The affected genes encode a wide range of apoptosis-related proteins, including death receptors (NTF-R1, NTF-R2, DR3, DR5, LTbeta-R, GITR, P57 TR-1) and their adaptor and regulatory proteins (MASGE-D1, TRAF-2, SIVA, MET, FLIP, FAIM, IAP1, ATFA), members of transcription regulatory pathways (JNK, NF-kappaB, P53), members of BCL-2 family of proteins (BID, BAD, BAX, BIK, NIP21, NIP3, NIX, BCL-2), DNA damage sensor (PARP-1), caspases and their substrates and regulatory proteins (caspases 8, 4, 9, and 3, ACINUS, CIDE-A, CIDE-B, GAS2), mitochondrially released factors (cytochrome c, AIF, PRG3), specific endoplasmic reticulum- and oxidative stress-associated factors (BACH2, ABL1, ALG2, CHOP), members of cell survival AKT and HSP70 pathways (PIK3GA, PTEN, HSP70, BAG1, BAG2), and others. This suggests that cocaine affects survival of developing cerebral cells via multiple apoptosis-regulating mechanisms. Experiment Overall Design: Animals Experiment Overall Design: Timed pregnant Swiss Webster (CFW; Charles River Experiment Overall Design: Lab. Wilmington, MA) dams were maintained in individual Experiment Overall Design: cages in a climate-controlled room on a 12/12-h light/ Experiment Overall Design: dark cycle. They were divided into two groups. The first Experiment Overall Design: (experimental) group received subcutaneous (at the dorsum Experiment Overall Design: of the neck) injections of 20 mg/kg cocaine hydrochloride Experiment Overall Design: (Research Technology Branch, National Institute Experiment Overall Design: of Drug Abuse, Rockville, MD) dissolved in 200 mkl of Experiment Overall Design: 0.9% saline, twice a day (at 8:00 AM and 8:00 PM) from Experiment Overall Design: 8th through 18th day of pregnancy (E8âE18). The second Experiment Overall Design: (control) group was subjected to the same schedule of Experiment Overall Design: 0.9% saline only injections. The cocaine treatment was Experiment Overall Design: designed to replicate the one capable of reducing cerebral Experiment Overall Design: cortical mass in mouse offspring. Also, the Experiment Overall Design: relatively protracted period the chronic treatment was Experiment Overall Design: chosen to maximize the changes in the tissue expression Experiment Overall Design: of cocaine-regulated genes of interest. Throughout the Experiment Overall Design: treatment, all mice were weighed daily, and, from E8, the Experiment Overall Design: control and experimental animals were pair-fed, with the Experiment Overall Design: daily amount of food (Mouse Chow; Ralston Purina Saint Experiment Overall Design: Louis, MO) provided to each control dam being matched Experiment Overall Design: to that consumed by the paired experimental dam. Water Experiment Overall Design: was available ad libitum. We found that this feeding Experiment Overall Design: regiment resulted in similar weight gains from E8 to E18 Experiment Overall Design: in both experimental and control animal groups Experiment Overall Design: (46.41F0.45% and 44.99F0.59% respectively). On E18, Experiment Overall Design: 1 h after the morning treatment, the animals were Experiment Overall Design: anesthetized by peritoneal injection of 50 mg/kg sodium Experiment Overall Design: pentobarbital (Abbott Lab., Abbott Park, IL) and their Experiment Overall Design: uteri were removed. The frontal cerebral wall (containing Experiment Overall Design: cerebral cortex and underlying transient cortical zones Experiment Overall Design: anterior to the striatal level) and the occipital cerebral wall Experiment Overall Design: (containing cerebral cortex and underlying transient Experiment Overall Design: cortical zones posterior to the level of the hippocampus) Experiment Overall Design: were dissected from the fetuses and stored in liquid Experiment Overall Design: nitrogen prior to analysis. The animal experimentations Experiment Overall Design: used in this study were approved by the University of Experiment Overall Design: Maryland Animal Care and Use Committee. Experiment Overall Design: Microarray Experiment Overall Design: Tissue from two control and two experimental fetuses Experiment Overall Design: were used in the processing of a single microarray slide Experiment Overall Design: (Part G4121A, Agilent Technol). Five slides were processed Experiment Overall Design: per region of the fetal cerebral wall. For a given region, the Experiment Overall Design: tissue for each microarray slide was obtained from a Experiment Overall Design: separate set of control and experimental litters. Total RNA Experiment Overall Design: was isolated using TRIzol Reagent (Invitrogen, Carlsbad, Experiment Overall Design: CA). The yield of total RNA was determined by absorbance Experiment Overall Design: at 260 nm on a DU 640 Spectrophotometer (Beckman Experiment Overall Design: Coulter, Fullerton, CA). The 260/280 nm ratios of the Experiment Overall Design: samples were >1.8. For each assay, 5 mkg of RNA from Experiment Overall Design: control tissue and 5 mkg of RNA from experimental tissue Experiment Overall Design: were reverse transcribed using 3DNA Array Detection Kit Experiment Overall Design: (Genesphere, Hatfield, PA) with primers containing different Experiment Overall Design: dcaptureT sequences for control and experimental Experiment Overall Design: samples. SuperScript II Reverse Transcriptase for these Experiment Overall Design: reactions was purchased from Gibco BRL (Gaithersburg, Experiment Overall Design: MD). The resultant cDNAs were hybridized to microarray Experiment Overall Design: slides for 16 h in Agilent Hybridization Buffer (Agilent Experiment Overall Design: Technol) at 60 8C. Upon completion of the hybridization, Experiment Overall Design: the slides were washed for 10 min with 0.005% Triton in Experiment Overall Design: 6xsodium chloride/sodium citrate buffer (pH 7; SSC) at Experiment Overall Design: room temperature and then for five more min with 0.005% Experiment Overall Design: Triton in 0.1xSSC and 1 more minute in 0.1xSSC (pH 7), Experiment Overall Design: both at 4 8C. Washed slides were air-dried for 30 s. For Experiment Overall Design: fluorescent labeling of microarray-bound cDNA, the slides Experiment Overall Design: were incubated for 3 h at 65 8C with Capture Reagent Experiment Overall Design: Hybridization Mixture from 3DNA Array Detection Kit Experiment Overall Design: (Genesphere). In this mixture, Alexa 546 fluorochromeincorporating Experiment Overall Design: dendrimers contained single-stranded arms Experiment Overall Design: complimentary to the dcaptureT sequences used in the Experiment Overall Design: reverse transcription of RNA from control tissue, while Experiment Overall Design: Alexa 647 fluorochrome-incorporating dendrimers contained Experiment Overall Design: arms complimentary to the capture sequences used Experiment Overall Design: in reverse transcription of RNA from experimental tissue. Experiment Overall Design: The labeling reaction was terminated by washing of the Experiment Overall Design: microarray slides at room temperature for 10 min with Experiment Overall Design: 0.005% Triton in 2xSSC and for another 10 min with in Experiment Overall Design: 0.2xSSC. After that, the slides were air-dried for 30 s. The Experiment Overall Design: processed microarray slides were scanned at 10 Am Experiment Overall Design: resolution on a GenePix 4100A scanner (Axon Instr., Union Experiment Overall Design: City, CA) with the laser excitation at 532 nm [emission filter Experiment Overall Design: 575DF35 (green); photomultiplier voltage 550] for Alexa Experiment Overall Design: 546 (control samples) and the laser excitation at 635 nm Experiment Overall Design: [emission filter 670DF40 (red); photomultiplier voltage Experiment Overall Design: 695] for Alexa 647 (experimental samples). The signals Experiment Overall Design: were converted into 16-bits-per pixel resolution images, Experiment Overall Design: providing color depth of 65,536 levels. The densitometry Experiment Overall Design: was performed with GenePix Pro 4.1 software (Axon Instr.). Experiment Overall Design: Background was subtracted using the dlocal background Experiment Overall Design: correctionT procedure available in the software. Quality control utilized 255 positive controls, 161 negative controls Experiment Overall Design: and 646 QC-spots present on Agilentâs arrays. For Experiment Overall Design: replicates within a slide, median signal values were Experiment Overall Design: calculated. For each array, the data were normalized in Experiment Overall Design: Acuity 3.1 software (Axon Instr.) by applying locally Experiment Overall Design: weighted scatterplot smoothing (LOWESS) transformation. Experiment Overall Design: The between array scale normalization Experiment Overall Design: was done using intensity log10 ratio distribution box plots (GeneSight Experiment Overall Design: software, BioDiscovery, El Segundo, CA) for verification. Experiment Overall Design: Statistical analysis of the normalized data was conducted Experiment Overall Design: by the Significant Analyses for Microarray Algorithm Experiment Overall Design: (SAM) using Excel macros available at the Stanford SAM website. Experiment Overall Design: In the analyses of both frontal and occipital regions of the fetal cerebral wall, Experiment Overall Design: the cut-off thresholds were set to identify the maximum Experiment Overall Design: number of cocaine exposure-regulated genes at the minimal Experiment Overall Design: false discovery rate (FDR) allowed by SAM for a given set Experiment Overall Design: of microarray data. For both regions, the FDR rates were Experiment Overall Design: <0.5%. The apoptosis and Wnt pathway-related genes were identified with Pathway Assist 2.0 software (Stratagene, La Jolla, CA). Experiment Overall Design: This was followed by manual review of literature for each Experiment Overall Design: identified gene.
2008-06-15 | E-GEOD-6628 | biostudies-arrayexpress
Project description:To understand hypoxia mediated changes in whole blood, normal C57Bl/10 mice were gradually exposed to a chronic chemical hypoxic environment, for 2 weeks. Control, age-machted mice were maintained under normoxic conditions. Purpose: To examine and characterize the expression profile of genes expressed at chemical hypoxia of whole blood in comparison to the control. Methods: At the beginning of the experiment mice were divided into two groups, control (room condition) and chemical hypoxic (room condition). For conditioning, the chemical hypoxic group was supplemented with 200ng of CoCl2 per 100ml/day. Animalâs weight and food intake were monitored daily. Food and water were changed daily during the course of the experiment. Before and after the experiments the hematocrit was monitored by taking blood from the tail vein of control and hypoxic animals. After 15 days animals were euthanized using CO2 after whole blood extraction from V. Cava for further analysis. RNA from whole blood was isolated, processed and used for microarray-based expression profiling. Profiles were generated for genes differentially expressed at control versus chemical hypoxia in whole blood using a false discovery rate (FDR) of 0%.We validated the profiles by real-time quantitative reverse transcription-polymerase chain reaction (qPCR). Results: The regional transcriptomes associated with chemical hypoxia in whole blood were identified. We found 3094 genes that were differentially expressed in chemical hypoxic whole blood compared to control with a 0% FDR and a 2 fold cutoff. Conclusion: Transcriptome level differences exist between control and chemical hypoxia in whole blood. Our definition of the synaptic transcriptome provides insight into the functioning of the unique response to hypoxia in whole blood.
2010-05-07 | E-GEOD-15902 | biostudies-arrayexpress
Project description:To determine hypoxia mediated changes in whole blood, normal swiss webster mice were gradually exposed to a chronic hypobaric hypoxic environment up to 8500m, for 2 weeks in vivo. Control, age-matched mice were maintained under normoxic conditions in Kathmandu (c. 1300 mts above sea level). Purpose: To examine and characterize the expression profile of genes expressed at hypobaric hypoxia on Mt. Everest of whole blood in comparison to the control. Methods: At the beginning of the experiment mice were divided into two groups, control (room condition, Kathmandu, Nepal) and hypoxic (hypoxic condition). For conditioning, the hypoxic group was exposed to lower levels of hypobaric hypoxia during our mountaineering expedition to Mt Everest. The oxygen level was decreased according to our climbing protocol from 21% to about 7% over a period of 15 days. Food and water were changed daily during the course of the experiment. After 15 days animals were euthanized after whole blood extraction from V. Cava for further analysis. RNA from whole blood was isolated, processed and used for microarray-based expression profiling. Profiles were generated for genes differentially expressed at control versus hypobaric hypoxia in whole blood using a false discovery rate (FDR) of 0%.We validated the profiles by real-time quantitative reverse transcription-polymerase chain reaction (qPCR). Results: The regional transcriptomes associated with hypobaric hypoxia on Mt. Everest in whole blood were identified. We found 947 genes that were differentially expressed in normobaric hypoxic whole blood compared to control with a 0% FDR and a 2 fold cutoff. Conclusion: Transcriptome level differences exist between control and hypobaric hypoxia in whole blood. Our definition of the synaptic transcriptome provides insight into the functioning of the unique response to hypoxia in whole blood.
2010-05-07 | E-GEOD-15901 | biostudies-arrayexpress
Project description:Purpose: To investigate the effect of transcorneal electrical stimulation (TES) on the retina of wildtype Brown Norway (BN) rats by gene expression profiling. Methods: TES was applied to BN adult wildtype rat retina in vivo for 1h (1ms biphasic pulses at 20Hz; current: 200 M-BM-5A). RNA was isolated, processed and used for microarray-based expression profiling 4hrs after initial TES. An expression profile was generated for genes differentially expressed at 4hrs after TES vs. sham stimulated animals using a fold change cutoff of 1.2. We validated the profile by real-time quantitative reverse transcription-polymerase chain reaction (qPCR). In addition, the application of TES was verified at the structural and functional level. Results: Transcriptome changes associated with TES vs. sham stimulated BN wildtype retina were identified. 490 genes were differentially expressed in TES and included well-known genes as well as a large number of novel genes. Electrophysiological recordings showed physiological retinal function after TES and structural in vivo and ex vivo studies revealed intact retinal layers. Conclusion: Our results demonstrate that TES applied to the retina of wildtype BN rats induce a variety of transcriptome level changes and may help to understand the mechanisms underlying TES. In addition TES has no negative effect on structure and function of wildtype BN retina 24hrs after application. RNA isolation and Microarray studies 4 animals were euthanized 4h after TES or respective sham treatment. Extraction of total RNA was performed using the RNeasy Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. QIAshredder mini-spin columns (Qiagen) as well as needle and syringe homogenization were applied for tissue homogenization. To ensure proper integrity all RNA used in our experiments showed single peaks for the 18S and 28S bands as determined by the Agilent 2100 Bioanalyzer using the RNA 6000 Nano LabChip Kit (both Agilent Technologies Inc., CA, USA) following the manufacturer's instructions. The purity and concentration of total RNA was determined by absorbance measurements at 260 and 280nm using a Nanodrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE, USA). All RNA that did not have a 260/280 ratio between 1.9 and 2.1 was discarded to ensure high quality of samples. Respective RNAs from stimulated and sham samples were used to generate double-stranded cDNA with the T7-oligo (dT) primer according to GeneChipM-BM-. (Affymetrix Inc., Santa Clara, CA, USA) two cycle target labeling protocol as described by the manufacturer. The processed cDNA was hybridized to Affymetrix GeneChipM-BM-. Rat Gene 1.0 ST arrays for 18M-bM-^@M-^S24h (Affymetrix Inc.). Four TES and four sham replicates were used for hybridization to Microarray GeneChipsM-BM-.. Each microarray was washed and stained with streptavidinM-bM-^@M-^Sphycoerythrin and scanned at a 6M-NM-<m resolution with Agilent Technologies model G2500A GeneArray scanner. A visual quality control measurement was performed to ensure proper hybridization after each chip was scanned. Additional quality control parameters such as scaling factors used to normalize the chips, average background, and noise were also evaluated. Microarray analysis For statistical analysis of microarray data all cell files were imported to GeneSpring GX software (Agilent Technologies Inc.) where GC-RMA algorithm was applied to yield log signal values on each probe set. All transcripts with a minimum change of 1.2 fold together with a p value less than 0.05 were selected using Significance Analysis of Microarray (SAM) (Tusher VG, 2001). The visualization of the relationships between the samples by principal component analysis (PCA) based on the GC-RMA signal and hierarchical clustering using the Pearson correlation was obtained using PARTEK Genomics Suite version 6.4. All differentially regulated genes were imported into Ingenuity Pathway Analysis (Ingenuity Systems, CA, USA) for generation of networks and canonical pathway analysis. The data set containing all significant gene identifiers along with corresponding expression and significance values was uploaded into the application. Each identifier was mapped to its corresponding object in Ingenuity's Knowledge Base. The set fold change cutoff identified all molecules whose expression was significantly differentially regulated. These network eligible molecules were overlaid onto a global molecular network developed from information contained in IngenuityM-bM-^@M-^Ys Knowledge Base. Networks were then algorithmically generated based on their connectivity. The functional analysis of a network identified the biological functions using a right-tailed FisherM-bM-^@M-^Ys exact test that were most significant to the molecules in the network. Canonical pathways analysis identified the pathways from the Ingenuity Pathways Analysis library of canonical pathways that were most significant to the data set. The significance of the association between the data set and the canonical pathway was measured in 2 ways: 1) A ratio of the number of molecules from the data set that map to the pathway divided by the total number of molecules that map to the canonical pathway is displayed. 2) FisherM-bM-^@M-^Ys exact test was used to calculate a p-value determining the probability that the association between the genes in the dataset and the canonical pathway is explained by chance alone.
2011-11-21 | E-GEOD-26174 | biostudies-arrayexpress
Project description:Analysis of the response to arginine of the Escherichia coli K-12 transcriptome by microarray hybridization and real-time quantitative PCR provides a first coherent quantitative picture of the ArgR-mediated repression of arginine biosynthesis and uptake genes. Transcriptional repression was shown to be the major control mechanism of the biosynthetic genes, leaving only limited room for additional transcriptional or post-transcriptional regulations. The art genes coding for the specific arginine uptake system are subject to ArgR-mediated repression, ; with a strong repression of artJ, coding for the periplasmic binding protein of the system. The hisJQMP genes of the histidine transporter (part of the LAO uptake system) were discovered to be a part of the arginine regulon. Analysis of their control region with reporter gene fusions and electrophoretic mobility shift in the presence of pure ArgR repressor showed the involvement in repression of the ArgR protein and of an ARG box 120 bp upstream of hisJ. No repression of the genes of the AO uptake system was observed. Finally, comparing the time course of arginine repression of gene transcription with the evolution of the specific ; activities of the cognate enzymes showed that while full genetic repression was achieved two minutes after arginine addition, enzyme concentrations were diluted at the rate of cell ; division. This emphasizes the importance of the feedback-inhibition of the first enzymatic ; step of the pathway in controlling the metabolic flow through the biosynthesis in the period following the onset of repression. Experiment Overall Design: 3 groups of 3 replicate samples were analyzed: Experiment Overall Design: (1) Wild type on minimal medium (strain P4X, slides 1753, 1765, 1989), this created a reference condition Experiment Overall Design: (2) Wild type grown in the presence of arginine (strain P4X plus 100µg/ml of arginine, slides 1754, 1766, 1990), this gave a list of all the genes repressed by the system: ArgR+arginine Experiment Overall Design: (3) Mutant where no active repressor is present; grown in the presence of arginine (strain P4XB2 (argR-) plus 100µg/ml arginine, slides 1992, 1993, 1994). This condition told us which genes are indeed regulated by the system: ArgR+arginine. Those genes are in this case derepressed.
2007-10-24 | E-GEOD-4724 | biostudies-arrayexpress