Steady State Thermal Acclimation in the Eurythermal Goby fish, Gillichthys mirabilis
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ABSTRACT: The goal of this study was to identify transcriptional resonses to thermal acclimation in a highly eurythermal fish. Specifically, what are the genes necessary to maintain homostasis at different temperatures and do these genes underlie any known acclimatory responses? Gillichthys mirabilis (goby fish) were acclimated to 3 temperatures (9C, 19C and 28C) for one month. Gill tissue from 7-9 individuals were dissected and immediately flash frozen in liquid nitrogen. Total RNA was extracted, reverse transcribed into cDNA, labeled with cy5 and hybridized against a common reference (cy3) on 2-color cDNA microarray developed for this species. Significant genes were determined by expression differences between groups (one-way ANOVA, FDR<0.05) . Common reference design
Project description:The goal of this study was to identify transcriptional resonses to acute heat stress following differential acclimation in the highly eurythermal goby fish, Gillichthys mirabilis. G. mirabilis were acclimated to 3 temperatures (9C, 19C and 28C) for one month and then exposed to an acute heat ramp at 4C/hr and sampled during the timecourse. Gill tissues were dissected and immediately flash frozen in liquid nitrogen. Total RNA was extracted, reverse transcribed into cDNA, labeled with cy5 and hybridized against a common reference (cy3) on 2-color cDNA microarray developed for this species. Significant genes were determined by expression differences between groups (one-way ANOVA, FDR<0.05) . Common reference design
Project description:Transcriptional analysis of the effects of natural environmental variation across the vertical distribution of Mytilus californianus within a single mussel bed Keywords: Environmental Response 30 Biological replicates from plots sampled at 3 different verticle tide heights above the MLLW at Strawberry Hill Oregon. 15 mussels were sampled after a mid-day emmersion event and 15 mussels were sampled after a 1 hour recovery at ambient seawater temperatures. 1 replicate per array, compared using a common reference sample. 50 Biological replicates for 5 plots sampled at 2 different verticle tide heights above the MLLW at Boiler Bay Oregon. 25 mussels were sampled after a mid-day emmersion event and 25 mussels were sampled after a 1 hour recovery at ambient seawater temperatures. Pooled RNA from 5 biological replicates from each plot per array, compared using a common reference sample.
Project description:Transcriptional analysis of the effects of natural environmental variation across the latitudinal range of Mytilus californianus 5 Biological replicates from 4 separate populations of mussels were compared using a common reference sample. Dye swap analysis was performed for each replicate.
Project description:Following an infection with a specific pathogen, the acquired immune system of many teleostean fish, including salmonids, is known to retain a specific memory of the infectious agent, which protects the host against subsequent infections. For example Atlantic salmon (Salmo salar), which have survived an infection with a low-virulence infectious salmon anemia virus (ISAV) isolate are less susceptible against subsequent infections with high-virulence ISAV isolates. A greater understanding of the mechanisms and immunological components involved in this acquired protection against ISAV is fundamental for the development of efficacious vaccines and treatments against this pathogen. To better understand the immunity components involved in this observed resistance, we have used an Atlantic salmon DNA microarray and RT-qPCR assays to study the global gene expression responses of preexposed Atlantic salmon (fish having survived an infection with a low-virulence ISAV isolate) during the course of a secondary infection with a high-virulence ISAV isolate Atlantic salmon which had survived a primary infection with a low-virulence ISAV isolate (preexposed H5R fish) were reinfected by cohabitation (H5Rc fish) or IP injection (H5Rip fish) with a high-virulence ISAV isolate and compared to preexposed H5R fish non-reinfected and naïve Atlantic salmon infected by cohabitation (Nc fish) with the high-virulence ISAV isolate using 4x44k Agilent arrays. H5Rc fish (n~6) and Nc fish (n~6) were sampled at 20d, 23d, 29d, 41d and 63d following the infection, while H5Rip fish (n~6) were sampled at 6h, 24h, 3d, 10d, 20d and 63d. Microarrays were performed using a 1-color approach
Project description:Long-term exposure of Atlantic salmon to 19M-BM-0C resulted in cardiac gene and protein expression changes indicating that the unfolded protein response, vascularization, remodeling of connective tissue and altered innate immune responses were part of the cardiac acclimation or response to elevated temperature Heart tissue from Atlantic salmon (RNA poooled from N=3 fish per 3 replicate tanks) reared under normal (14 deg C) and elevated (19 deg C) temperatures were sampled after 21 (short-term) and 56 days (long-term acclimation), and gene expression changes between the elevated and the control temperature were assessed using microarray analysis.
Project description:We analyzed global transcriptional changes in both shoots and roots of root-flooded Arabidopsis seedlings by microarrays. We also interpreted the significance of the systemic communication between roots and shoots by functional classification of affected genes. We performed genetic analysis with an ethylene signaling mutant, ein2-5, to correlate systemic flooding responses with ethylene signaling. We identified a class of genes that were up- or downregulated in shoots, but not affected in roots, under hypoxic conditions. A comprehensive managing program of carbohydrate metabolism was observed, providing an example of how systemic communications might facilitate the survival of plants under flooding. A proportion of long-distance hypoxic regulation was altered in ein2-5. Time course experiments (0.5, 1, 3, 6, and 12h for Columbia; 0.5, 3, and 6h for ein2-5). Tissues from root-flooded seedlings vs. Tissues from un-flooded seedlings. Biological replicates: 4 replicates for each time point, independently grown, treated, and harvested. One replicate per array. 2 of 4 replicates are dye-swapped.
Project description:Light is a universal environmental signal perceived by many organisms, including the fungi in which light regulates both common and unique biological processes depending on the species. We conducted a whole-genome microarray analysis on the basidiomycete fungus Cryptococcus neoformans to identify light-regulated genes. Two-condition experiment, cells grown in complete darkness or exposed to white light. Six biological replicates independently grown and harvested. One replicate per array.
Project description:Atlantic cod (~70 g) were exposed to water temperatures of 12 and 17 degrees Celsius (flow-through system), then half the fish in each temperature group were infected with Francisella noatunensis (injection). Fish were sampled prior to temperature challenge, when water temp reached 17 degrees, and five weeks after infection. The left section of the olfactory organ was collected on RNAlater for RNA isolation and sequencing.
Project description:To determine the effects on the intestine gene expression of pathogen exposure to Enteromyxum leei. One fish group was exposed to E. leei-contaminated effluent (recipient group = R) . R fish (n= 66, average weight = 134 g) were placed in two replicated 200L fibre-glass tanks which were set to receive exclusively the effluent water from another tank containing 24 infected (donors = D; average weight = 127.3 g; prevalence of infection = 54%) gilthead sea bream. The D to R fish ratio was 0.8. Other 66 naïve fish were allocated in two replicated tanks (control group = CTRL) under the same conditions, but without receiving contaminated effluent. Over the course of the study, day length followed natural changes and water was heated in order to keep temperature always above 18ºC, the range was 18-23 ºC. Water was 5 µm-filtered and UV irradiated, and salinity was 37.5â°. Water flow was 10L/min and oxygen content of outlet water remained higher than 85%saturation. All fish were fed daily a commercial dry pellet diet at about 1% of body weight. Disease signs and daily mortalities were recorded throughout the experiments. The parasitic status of dead fish was checked by microscopic examination of fresh intestinal scrapings. Fish were sampled after 113 days post exposure (p.e.). Feeding was stopped one day prior to the sampling to ensure that the digestive tract was empty. Head kidney and posterior intestine were rapidly excised, frozen in liquid nitrogen, and stored at -80 °C until RNA extraction and analysis. Keywords: Treated/Untreated, confinement, cortisol, stress response, time course, microarray 28 intestine samples - Twenty eight slides were hybridised using a reference design. Three groups (control, infected and non-infected) were compared using five individual fish in each group. Each sample was hybridised twice - the second being a dye-swap of the first.
Project description:The unicellular cyanobacterium Synechocystis sp. PCC 6803 is a model system for studying biochemistry, genetics and molecular biology of photobiological processes. Despite its importance in basic and applied research, the genome-wide picture of transcriptional regulation in this bacterium is limited. Characteristic transcriptional responses to changes in the growth environment are expected to provide a scaffold for describing the Synechocystis transcriptional regulatory network as well as efficient means for functional annotation of genes in the genome. We designed, validated and used Synechocystis genome-wide oligonucleotide (70-mer) microarray (representing 96.7% of all chromosomal ORFs) to study transcriptional activity of the cyanobacterial genome in response to S deprivation. The microarray data were verified by quantitative RT-PCR. We made five main observations: 1) Transcriptional changes upon sulfate withdrawal were relatively moderate, but significant and consistent with growth kinetics; 2) S acquisition genes encoding for a high-affinity sulfate transporter were significantly induced, while decreased transcription of genes for phycobilisome, photosystems I and II, cytochrome b6/f, and ATP synthase indicated reduced light-harvesting and photosynthetic activity; 3) S deprivation elicited transcriptional responses associated with general growth arrest and stress; 4) A large number of genes regulated by S availability encode hypothetical proteins or proteins of unknown function; 5) Hydrogenase structural and maturation accessory genes were not identified as differentially expressed, even though increased hydrogen evolution was observed. The expression profiles recorded by using this oligonucleotide-based microarray platform revealed that during transition from the condition of plentiful sulfur to no sulfur, Synechocystis undergoes coordinated transcriptional changes, including genes whose products are involved in sensing nutrient limitations and tuning bacterial metabolism. The transcriptional profile of the nutrient limitation was dominated by decrease in abundances of many transcripts. However, these changes were unlikely due to the across-the-board, non-specific shut down of transcription in a condition of growth arrest. Down-regulation of transcripts encoding proteins whose function depends on a cellular sulfur status indicated that the observed repression has a specific regulatory component. The repression of certain sulfur-related genes was paralleled by activation of genes involved in internal and external S scavenging. Keywords: stress response, time course Synechocystis sp. PCC 6803 was grown photoautotrophically in BG-11 medium supplemented with 8mM NaHCO3 and buffered with 10mM HEPES (pH 7.4). The cells were grown in 250ml flasks at 32oC under a light intensity of 25µmol photons m-2 s-1. Cultures were bubbled with sterile air containing 1% (v/v) CO2. Log phase cells (OD730nm=0.6) were harvested by centrifugation (2000×g for 12 min) washed once and then re-suspended in sulfate-free media (MgSO4 replaced by the same molarity of MgCl2). In addition, all S-containing trace metals in BG-11 were replaced by non-S containing metals. Cells were harvested and fixed for microarray analysis by adding 10% (v/v) ice-cold 5% phenol in ethanol stop solution at the following time points: before S-depravation (time 0, control), 1, 3, 6, 12, 24, 48 and 72 hr after S-depravation. S-deprivation with HEPES buffering control experiment was performed as described above, except that HEPES buffer was used upon sulfate removal. Bacterial samples for a time course were taken at time 0, 1, 12 and 24 hrs after sulfate withdrawal. Growth stage control experiment was done in parallel with S deprivation experiments. Samples were taken at 0, 1, 2.5, 4, 7, 11 and 48 hr after OD730nm reached 0.60. All the experiments were done in biological replicates.