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Sample GSM948767 Query DataSets for GSM948767
Status Public on Jun 15, 2012
Title Brain from Gaighatta Female rep 2
Sample type RNA
 
Source name Brain from wild Gaighatta strain, Univ. of Idaho
Organism Danio rerio
Characteristics strain: Gaighatta
tissue: Brain
rearing history: Wild
age: Adult
Sex: Female
Treatment protocol Sixteen adult fish (4 to 6 months old) from each strain were randomly assigned to 1-L aquaria (22 cm long, 15 cm high, 5 cm wide), one fish per aquarium. The aquaria were delimited into six 2-cm vertical zones using thin twine tied across the front of the entire row of aquaria. Fish were then observed using a high-throughput behavioral assay to quantity fear-related behaviors. Following behavioral observations, fish were removed from the individual tanks and temporily placed in one of sixteen 3-L group tanks. On the sampling day, each fish was anesthetized in 170 mg L−1 tricaine methanesulfonate (MS222, Western Chemical Inc., Ferndale, WA), briefly blotted on a paper towel, and rapidly measured for standard length and body mass. The brain was then quickly removed and homogenized in TRIzol (Invitrogen, Carlsbad, CA). Brains from all individuals in a tank were homogenized together, for a total of two biological replicate pools per sex per strain (16 microarrays total).
Growth protocol Prior to the experiment, all zebrafish were reared in a recirculating zebrafish facility at the University of Idaho, designed by Aquaneering Inc. (San Diego, CA). Fish were feed twice daily with a combination of commercial flake food and Artemia nauplii. Temperature of the facility was maintained at 28ºC with a constant 14 h light: 10 h dark cycle. These rearing conditions were maintained during the behavior assays.
Extracted molecule total RNA
Extraction protocol Total RNA was then extracted using the TRIzol method following manufacturer’s protocol (Invitrogen).
Label biotin
Label protocol For each array, 10 ug of total RNA were converted to cDNA. Biotinylated cRNA was then produced in vitro using the GeneChip expression 3′ amplification one-cycle target labeling kit (Affymetrix, Santa Clara, CA, USA).
 
Hybridization protocol Affymetrix Zebrafish Genome Arrays (~14,900 transcripts) were hybridized with fragmented biotinylated cRNA for 16 h at 45°C with constant rotation (45 rpm), and processed using the Affymetrix GeneChip Fluidic Station 450. Streptavidin-conjugated phycoerythrin (SAPE) was used for staining, followed by amplification using a biotinylated anti-streptavidin antibody. This was followed by another round of SAPE. All microarray procedures were performed at the Genomics Core Facility of the Center for Reproductive Biology at Washington State University (Pullman, WA).
Scan protocol All microarrays were scanned using a GeneChip Scanner 3000 (Affymetrix).
Description Gene expression data from brain of wild Gaighatta zebrafish strain
Data processing CEL files containing raw data were then processed and analyzed using R software and Bioconductor packages. Microarray hybridization data were examined for physical anomalies on the chip by pseudochip and residual error visualizations. Quality assurance of microarray data was completed using the affyQAReport function from the Bioconductor package affyQCReport. The arrays were then pre-processed using the Robust Multi-array Average (RMA) procedure using the affy package. Next, unexpressed and low variability genes were removed by unbiased filtering. Affymetrix present-marginal-absent (PMA) calls were determined using a P-value cut off for absent of greater than 0.04 and present less than 0.04; marginal calls were treated as absent. Unexpressed genes were then defined as having a signal less than the expression value at which 99% of genes were called as absent across all samples. A filter on interquartile range was also applied to remove genes with low variability. Genes with an interquartile range of less than 0.5 across all chips in the experiment were excluded, reducing the dataset further to 7,958 genes. Signal intensities were also examined at the probe level to identify single feature polymorphisms (SFPs), differences at the probe level due to genetic polymorphisms rather than expression differences, which may also impact computed expression values. Briefly, in R using previously described methods, the RMA normalized expression estimate for each probe set was subtracted from background corrected and normalized expression levels at individual probes within the probe set. Normalized residuals were analyzed using significance analysis of microarrays (SAM) within the siggenes package to detect features with a significant effect for strain (FDR adjusted < 0.01). A total of 3,199 genes with significant SFPs were then removed from the analysis.
 
Submission date Jun 14, 2012
Last update date Jun 15, 2012
Contact name Robert Drew
E-mail(s) rdrew@umassd.edu
Phone 508-990-8950
Organization name University of Massachusetts Dartmouth
Department Biology
Street address 285 Old Westport Road
City N. Dartmouth
State/province MA
ZIP/Postal code 02747
Country USA
 
Platform ID GPL1319
Series (1)
GSE38729 Brain transcriptome variation among behaviorally distinct strains of zebrafish (Danio rerio)

Data table header descriptions
ID_REF
VALUE RMA Normalized Signal Intensity

Data table
ID_REF VALUE
AFFX-Dr-M62653-1_at 10.15369501
AFFX-Dr-M62653-1_s_at 8.946407222
AFFX-Dr-AF292560-1_s_at 9.216951467
Dr.3374.1.S1_at 4.109209562
Dr.19471.1.A1_at 6.034569546
Dr.13609.1.A1_at 8.263091948
Dr.13531.1.A1_at 9.427269998
Dr.13609.2.S1_at 8.655138204
Dr.9478.1.S1_at 5.151656182
Dr.23469.1.S1_s_at 3.323974122
Dr.9117.1.A1_at 8.057530849
Dr.25322.1.S1_at 4.27074536
Dr.10066.1.A1_at 2.567041323
Dr.26538.1.A1_at 2.913689335
Dr.5569.2.A1_at 7.444754939
Dr.4198.1.S1_at 3.164617948
DrAffx.2.21.S1_at 6.009749286
Dr.5853.1.A1_at 5.655551385
Dr.16385.1.A1_at 7.769621635
Dr.5000.1.A1_at 7.33175891

Total number of rows: 7958

Table truncated, full table size 222 Kbytes.




Supplementary file Size Download File type/resource
GSM948767_Robison-16-G-020706.CEL.gz 1.9 Mb (ftp)(http) CEL
Processed data included within Sample table

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