P. vivax: proof-of-principle global transcription analysis from two wild isolates
Ontology highlight
ABSTRACT:
ORGANISM(S): Plasmodium vivax
SUBMITTER: Hernando del Portillo
PROVIDER: E-GEOD-11075 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:The existence of two separate lineages of Escherichia coli O157:H7 has previously been reported, and research indicates that one of these lineages (lineage I) might be more pathogenic towards human hosts. We have previously shown that the more pathogenic lineage expresses higher levels of Shiga toxin 2 (Stx2) than the non-pathogenic lineage II. To evaluate why lineage 2 isolates do not express appreciable levels of toxin, two lineage 2 isolates (FRIK966 and FRIK2000) were chosen as representatives of lineage 2 and whole genome microarrays were performed using Agilent microarrays using the E. coli O157:H7 EDL933 lineage I clinical type isolate as a reference. Microarray results were utilized to evaluate what genes and pathways might be missing or differentially expressed. Quantitative RT-PCR was utilized to validate the microarray data. Based upon the transcriptome of Escherichia coli O157:H7 EDL933 an oligonucleotide microarray, made up of 60 mers was designed. A total of 4873 genes in an 8 x 15K Agilent microarray design. Designed using a custom script, specifications for gene specific oligos were based upon various design characteristics such as temperature of melting, 3’ location, specificity, lack of repeat nucleotides, etc. (Charbonnier et al., 2005). Arrays were manufactured using Agilent Sure-print technology. Each array consisted of duplicate elements for each gene randomly distributed with Agilent control elements included. All procedures were performed according to respective manufacturer protocols. Lineage I and lineage II strains were grown overnight as described, a total of 10e7 cells were washed twice in fresh media, normalized based upon optical density, inoculated into fresh media, incubated at 37oC shaking 120 x g for 3 hours and then suspended in RNAprotect bacteria reagent (Qiagen Inc., Valencia, CA.). Total RNA was extracted using RNeasy Bacteria Mini Kit (Qiagen Inc., Valencia, CA.) and trace amounts of DNA were removed using RNase-Free DNase Set (Qiagen Inc., Valencia, CA.). RNA was quantified using Nanodrop system (NanoDrop Technologies, Wilmington, DE) and quality confirmed by electrophoresis on a Bio-rad Experion system (BioRad XXX). For each sample, 10 ug of RNA were labeled with either CyDye3-dCTP or CyDye5-dCTP (Perkin Elmer) using the LabelStar kit (Qiagen Inc., Valencia, CA.) and Random nonamers (Integrated DNA Technologies ). Labeled cDNA were hybridized to the microarray using Agilent Hi-RPM hybridization solution in an Agilent Hybridization chamber (Agilent.). A total of 8 arrays, each with duplicate elements for each gene, alternating dye swap for each replicate (4 biological replicates), were analyzed to obtain genes that were consistently and differentially regulated in comparison to EDL933, while limiting false discover rate (FDR) below a stringent 5% (Benjamini and Hochberg, 1995).
2009-08-31 | E-GEOD-17070 | biostudies-arrayexpress
Project description:Anopheles gambiae mosquitoes exhibit an endophilic, nocturnal blood feeding behavior. Despite the importance of light as a regulator of malaria transmission, our knowledge on the molecular interactions between environmental cues, the circadian oscillators and the host seeking and feeding systems of the Anopheles mosquitoes is limited. In the present study, we show that the blood feeding behavior of mosquitoes is under circadian control and can be modulated by light pulses, both in a clock dependent and in an independent manner. Short light pulses (~2-5 min) in the dark phase can inhibit the blood-feeding propensity of mosquitoes momentarily in a clock independent manner, while longer durations of light stimulation (~1-2 h) can induce a phase advance in blood-feeding propensity in a clock dependent manner. The temporary feeding inhibition after short light pulses may reflect a masking effect of light, an unknown mechanism which is known to superimpose on the true circadian regulation. Nonetheless, the shorter light pulses resulted in the differential regulation of a variety of genes including those implicated in the circadian control, suggesting that light induced masking effects also involve clock components. Light pulses (both short and longer) also regulated genes implicated in feeding as well as different physiological processes like metabolism, transport, immunity and protease digestions. RNAi-mediated gene silencing assays of the light pulse regulated circadian factors timeless, cryptochrome and three takeout homologues significantly up-regulated the mosquito's blood-feeding propensity. In contrast, gene silencing of light pulse regulated olfactory factors down-regulated the mosquito's propensity to Our study suggests that the mosquitoâs feeding behavior is under circadian control. Long and short light pulses can induce inhibition of blood-feeding through circadian and unknown mechanisms, respectively, that involve chemosensory factors. A series of assays were performed to assess transcriptomic changes in mosquitoes upon light stimulation and blood feeding in order to assess relationships between photic stimulation and modulation of feeding behavior.
2010-06-24 | E-GEOD-13560 | biostudies-arrayexpress
Project description:Background: The mosquito Anopheles gambiae is a major vector of human malaria. Increasing evidence indicates that blood cells (hemocytes) comprise an essential arm of the mosquito innate immune response against both bacteria and malaria parasites. To further characterize the role of hemocytes in mosquito immunity, we undertook the first genome-wide transcriptomic analyses of adult female An. gambiae hemocytes following infection by two species of bacteria and a malaria parasite. Results: We identified 4047 genes expressed in hemocytes, using An. gambiae genome-wide microarrays. While 279 transcripts were significantly enriched in hemocytes relative to whole adult female mosquitoes, 959 transcripts exhibited immune challenge-related regulation. The global transcriptomic responses of hemocytes to challenge with different species of bacteria and/or different stages of malaria parasite infection revealed discrete, minimally overlapping, pathogen-specific signatures of infection-responsive gene expression; 105 of these represented putative immunity-related genes including anti-Plasmodium factors. Of particular interest was the specific co-regulation of various members of the Imd and JNK immune signaling pathways during malaria parasite invasion of the mosquito midgut epithelium. Conclusion: Our genome-wide transcriptomic analysis of adult mosquito hemocytes reveals pathogen-specific signatures of gene regulation and identifies several novel candidate genes for future functional studies. In order to identify hemocyte-specific and immune-responsive transcripts, we first compared transcripts expressed in hemocytes from one day old sugar-fed mosquitoes to transcripts detected in whole mosquitoes of the same age and feeding status. This resulted in identification of the hemocyte-enriched transcriptome. We then compared hemocytes from 1 day old mosquitoes, 1 hour after immune challenge with heat-killed Escherichia coli or Micrococcus luteus, to control female mosquitoes injected with sterile PBS to determine the bacteria challenge responsive transcriptomes. We used heat-killed bacteria in these assays, because our primary interest was in identifying the bacterial responsive transcriptome and to avoid the potentially confounding effects of altered gene expression due to the lethal effects of a systemic infection associated with injection of living bacteria. Lastly, we compared hemocytes from mosquitoes at 24 hours and 19 days after ingestion of a blood meal infected with Plasmodium berghei to mosquitoes of the same age fed a non-infected blood meal to determine the ookinete and sporozoite infection responsive transcriptomes, respectively. This design resulted in a total of five experimental treatments. The following samples are not included in this submission: Hemo E coli vs. hemo unchallenged A Hemo E coli vs. hemo unchallenged B Hemo m luteus vs. hemo unchallenged A Hemo m luteus vs. hemo unchallenged B
2011-10-31 | E-GEOD-33269 | biostudies-arrayexpress
Project description:The local background was subtracted from the fluorescent value of each spot. Feature intensities were extracted from scanned microarray images using GenePix Pro 5.1. (Axon Instruments). The images were visually inspected, and spots from low-quality areas of the array were flagged and excluded from further analysis. Spots were also excluded from analysis if both the combined fluorescent intensity for both channels was less than 1.4 times that of the local background and the pixel-by-pixel correlation coefficient of the spot was less than 0.4. The fluorescence ratios were normalized as previously described (Turton and Gant oncogene 2001website). A hierarchical clustering algorithm (M. Eisen Brown and Botstein PNAS 98 website) was applied to those genes that fulfilled all of the following conditions: they were not flagged; they were differentially expressed (equal or higher than 1.5 fold upregulated or equal or lower than 2 fold downregulated); they had p-values equal or less than 0.02; and they were consistent on at least 4 out of the 5 arrays.
2010-05-27 | E-GEOD-2069 | biostudies-arrayexpress
Project description:A set of 22 expts. aimed at identifying splicing events dependent upon on the Spt4-5 transcription elongation factors in yeast. Four spt mutants and an mRNA capping mutant were analyzed four times each, including biological and technical (dye-swap) replicates. Two wt vs wt expts. were also performed. Keywords = transcription/spt5/spt4/splicing/DEDS
2010-06-09 | E-GEOD-1610 | biostudies-arrayexpress
Project description:The goal of this study is to identify P. vivax genes whose expression is dependent on the intact spleen in experimental infections in Aotus monkeys. These studies were carried-out at the facilities of the “Fundación Centro de Primates de la Universidad del Valle”, Cali, Colombia and in the “Barcelona Centre for International Health Resarch” - CRESIB, Barcelona, Spain. This protocol was approved from the Ethical Committees of both Centres. A total of 4 Aotus lemurinus griseimembra naive animals were used in these experiments. Three animals were splenectomized whereas another had an intact spleen. A donor monkey was infected with P. vivax Sal-I strain and after peak parasitemias appeared a time-series infections into Sp-1, Sp-2, Sp-3, and Sp+2 animals were performed. Parasites from each infection were obtained from peripheral blood, monkey leukocytes were removed by MidMacs and only purified schizont stages were used for RNA extractions. Dual-hybridizations Cy3/Cy5 comparing the global expression of parasites obtained from different infections (Cy5) with a reference pool PvSp-1 obtained from splenectomized monkeys from CDC (PvSp-1) were perfomed using an Agilent's 60-mer platform representing the complete coding genome of P. vivax (1 oligonucleotide/2 kb coding sequences) GPL6667
2011-07-10 | E-GEOD-12174 | biostudies-arrayexpress
Project description:This series represents the effects of OVA, tg-IL-13, and direct effects of IL-13 on airway epithelial cells. Experiment Design: Type of experiment: comparison of three related murine models of asthma, 1) Ova model, 2) tg-IL-13 model, 3) IL-13/Epi model. Two control groups were used: PBS challenged mice to control for the effects of OVA and tg-IL-13+ and STAT6-/- mice as controls for tg-IL-13 and IL-13/Epi mice. Measurements were made in each model in two types of tissue samples, whole lung and tracheal perfusate. Experimental factors: Allergen vs sham challenge, natural STAT6 expression vs transgenic expression and no expression. The number of hybridizations performed in the experiment: 50 (25 whole lung and 25 tracheal perfusate). The type of reference used for the hybridizations, if any: Pooled reference from lung (whole lung or tracheal perfusate) from untreated wildtype mice. Hybridization design: two-color hybridizations with reference design. Each individual sample was hybridized to a separate array. Quality control steps taken: 5 experimental replicates per group. RNA integrity assessed by Agilent Bioanalyzer. Arrays with low signals, high background, or high spatial variation rejected and corresponding samples reanalyzed. URL of any supplemental websites or database accession numbers:GEO (http://www.ncbi.nlm.nih.gov/geo, accession number GSE1438). ********** Samples used, extract preparation and labeling: The origin of the biological sample: homogenized whole lung from mouse and perfusate of mouse trachea. Manipulation of biological samples and protocols used: Whole lungs mechanically homogenized in 7.0 ml Trizol reagent (Invitrogen) for total RNA collection. Tracheas perfused with lysis buffer (Qiagen Rneasy kit) for total RNA collection. Protocol for preparing the hybridization extract and labeling: Preparation of aminoallyl-UTP labeled whole lung cDNA was performed as described [1]. Total RNA from tracheal perfusate (1.0 - 1.5 g per sample) was amplified and labeled with aminoallyl-dUTP using the MessageAmp aRNA kit (Ambion). Labeled cRNAs were coupled to Cy3 or Cy5 dyes (CyScribe, Amersham Biosciences) and purified as described [1]. Labeling protocol(s): Coupled to Cy3 or Cy5.External controls (spikes): none. ********** Hybridization procedures and parameters: Hybridizations were performed as described [1] except Ambion SlideHyb Glass Array Hybridization Buffer #1 (neat concentration) was used as hybridization buffer and hybridizations were carried out for 40 h at 55 °C. Following hybridization, arrays were washed in 1X SSC with 0.03% SDS (wash 1), 0.2X SSC (wash 2), and 0.05X SSC (wash 3) for five minutes for each wash. ********** Measurement data and specifications: Arrays were scanned using an Axon Genepix 4000B scanner and GenePix Pro Analysis 5.0 software; laser power 100%, 10 micron resolution, PMT optimized for each array. Data from GPR files are available from GEO (http://www.ncbi.nlm.nih.gov/geo, accession number GSE1438).The âprint-tip loessâ normalization was used to correct for within-array dye and spatial effects and single channel quantile normalization was used to facilitate comparison between arrays. No background subtraction was performed. We used functions in the library marrayNorm of the R / Bioconductor package to perform these normalizations. After normalization we determined the log2 ratio of experimental sample intensity to reference sample intensity for each probe on each array. ********** Array Design: General array design Array design name: UCSF 10Mm Mouse v.2 Oligo Array (GEO GPL1089) and UCSF Gladstone 18K Mouse v.2 Oligo Array (GPL1196) Platform type: spotted oligonucleotides (70mers) Surface and coating specification: aminosilane coated glass slides Physical dimensions of array support (e.g. of slide): 25 x 75 x 1.0 mm Number of features on the array: GPL1089: 19152, GPL1196: 18240 (including empty and duplicate features) For production protocol, see http://www.microarrays.org/pdfs/PrintingArrays.pdf Feature information is available from GEO (GSE1438). Reporters: synthetic single stranded oligonucleotides. Sequence and annotation information available from GEO (GPL1089 and GPL1196) Method of reporter preparation: synthesized by Operon. The spotting protocols used, including the array substrate, the spotting buffer, and any post-printing processing, including cross-linking: see http://www.microarrays.org/pdfs/PrintingArrays.pdfAny additional treatment performed prior to hybridization: none. ********** Reference: 1. Barczak A, Rodriguez MW, Hanspers K, Koth LL, Tai YC, et al. (2003) Spotted long oligonucleotide arrays for human gene expression analysis. Genome Res 13: 1775-1785. Keywords = IL-13 Keywords = Epithelial Keywords = differential gene expression Keywords = microarray
2010-06-10 | E-GEOD-1438 | biostudies-arrayexpress
Project description:Analysis of Culex quinquefasciatus responses to West Nile virus (WNV) infection at 7 and 14 days after ingestion of infected blood in the gut and carcass tissues. Comparison of WNV-infected to non-infected carcass and gut samples.
2011-11-01 | E-GEOD-23045 | biostudies-arrayexpress
Project description:We used cDNA microarray technology to compare the genome-wide expression profiles of a wild type strain (BY4700) (E02, E04, E06) or the isogenic strain deleted of GLN3 and GAT1 genes (E01, E03, E05) grown in YNB medium with glutamine as nitrogen source (M.Gln) against the wild type strain grown in M.Gln after addition of rapamacyn (20 min) (E01, E02), or M.proline (M.Pro) (E03, E04) or after a two hours shift from M.Gln to M.Pro (E05, E06), all growth conditions known to modify the expression of genes involved in nitrogen utilization. These microarrays allowed to identify the set of genes that were up or down-regulated in response to the quality of the nitrogen source. To evaluate whether the majority of genes responding to the nitrogen source were dependent on Gln3 and Gat1, we compared the expression profiles of the wild type strain and of the isogenic strain deleted of GLN3 and GAT1 genes (03167b: ura3, gln3â, gat1â), when both strains were grown on M.Gln + rapamycin (E07), or M.Pro (E08) or after a shift from M.Gln to M.Pro (E09). We also used an independent means of identifying Gln3-Gat1 regulated genes by comparing the expression profiles in wild type and ure2â (4709âURE2) strains on M.Gln medium (E10). The hybridization signal was measured using a GSM418 laser scanner. Image analysis for each array was processed using the GenePix Pro 4.0 (Axon Instruments, Inc.) software package, which measures fluorescence intensity pairs for each gene. Following image acquisition, a visual inspection of the individual spots on each microarray (size, signal-to-noise ratio, background level, and spot uniformity) completed the flagging (present/not present, good/bad) of the data. To maximize sensitivity two scans were made, one at high laser power and high PMT (Photo Multiplier Tube) gain to detect the faintest spots, and a second one using low laser power and a PMT gain avoiding saturation. The values for spots presenting ⥠5% saturation in the first scan were calculated based on an extrapolation after linear regression analysis of the intensities from both scans. These data were then imported in the GeneSpring 7.1 (Silicon Genetics) software package, applying a per spot per chip intensity-dependent (Lowess) normalization for further analysis.
2010-07-01 | E-GEOD-2891 | biostudies-arrayexpress
Project description:all data from control and diabetic
2010-06-05 | E-GEOD-2068 | biostudies-arrayexpress