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SEA BREAM_CONFINEMENT

ABSTRACT: Juvenile gilthead sea bream of 10-15 g initial body weight were randomly distributed in eight 500-L tanks in a seawater re-circulatory system equipped with physical and biological filters, and a heat-unit system that maintained water temperature above 18-19 ºC. Fish grew from July to December at a density of 8-10 kg/m3 with an overall daily growth index of 1.9 ± 0.01. Extruded pellets (47 % protein, 21% lipid; Proaqua, Palencia, Spain) were offered to visual satiety. Feeding was stopped one day before confinement exposure, and batches of 10 fish were then transferred from two 500-L tanks to cylinder net baskets of 10-L volume (117-123 Kg/m3), suspended each one in 90-L tanks with a high flow of seawater (10 L/min) to avoid water deterioration (Oxygen > 5 ppm; unionised ammonia < 0.02 mg/L). Fish from seven additional 500-L tanks were established as control groups, and remained undisturbed prior to sampling at zero and each sampling time. At each sampling time (1.5, 3, 6, 24, 72 and 120 h), eight fish from one control and one confinement tank were netted into a bucket to be anaesthetized with 0.1 g/L 3-aminobenzoic acid ethyl ester (MS-222). Fish were killed by cervical section and liver was taken, frozen immediately in liquid nitrogen and stored at -80 ºC until RNA isolation. Only 6h, 72h and 120h were analysed on the microarray. The experimental design was a reference design with the reference being prepared from a pool of all (confined and control) RNAs used in the microarray analysis. All samples were hybridised twice (except L72C5R, L120C2F, L120S4F, L24C4F) with one being the dye swap of the other. Keywords: Time Course

ORGANISM(S): Sparus aurata

PROVIDER: GSE16633 | GEO | 2009/12/31

SECONDARY ACCESSION(S): PRJNA117349

REPOSITORIES: GEO

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Project description:Gene expression analyses have been performed on brain tissue of sexually mature and immature males using microarrays. 60 eels were transferred to two independent temperature controlled recirculation water units connected to two 500 L cylindro-conical tanks (30 fish per tank) where the fish were acclimated to seawater (35 PSU salinity) over a 2 week period.Eel males in one of the seawater units were injected intramuscularly every week over a 140 day period with 2000 IU hCG/kg (human chorionic gonadotropin, Sigma–Aldrich Chemical) dissolved in 0.9% saline to induce sexual maturation. Eel males in the other recirculation unit were injected weekly over the same period with 0.9% NaCl (vehicle).At the end of the experiment, eels were anesthetized in a solution of 0.1mg/L tricaine methanesulfonate (MS-222, Sigma Aldrich) and blood samples collected into heparinized syringes. Tissues (brain, including the olfactory bulbs, telencephalon, diencephalon and mesencephalon, and gonads) were collected from sexually immature (n=12) and sexually mature males (n=12).RNA was extracted from brain tissue of a total of 24 (12 mature and 12 immature) males. For each individual, RNA was extracted separately for olfactory bulb, telencephalon and diencephalon. Because RNA concentrations of olfactory bulb, telencephalon and diencephalon were below the amount needed for microarray analysis, a separate experiment for each part of the brain was not possible. Therefore, we combined equal concentrations of olfactory bulb, telencephalon and diencephalon as a single brain sample for each individual.Microarray analysis was conducted using an European eel-specific array consisting of a total of 14,913 probes based on a large collection of high-throughput transcriptomic sequences (Pujolar et al. 2012). Probe sequences and further details on the microarray platform can be found on the GEO database under accession number GPL15124. Data was normalized using a quantile normalization procedure using R (http://www.r-project.org)

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SEA BREAM_CONFINEMENT

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