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Sample GSM323510 Query DataSets for GSM323510
Status Public on Apr 15, 2010
Title oocyte, biological rep 3
Sample type RNA
 
Source name Bovine oocyte
Organism Bos taurus
Characteristics Age: oocyte
Treatment protocol As described in growth protocol, no other treatment applied.
Growth protocol Ovaries were washed and 2-6 mm follicles were aspirated using an 18gauge needle. Only oocytes surrounded by several cell layers of dense cumulus were selected for culture. Oocytes were washed three times in TL-HEPES (0.22mM sodium pyruvate, 3mg/ml BSA, pH 7.4) and placed 10 per 50 ml drop of maturation media that had been equilibrated in 5% carbon dioxide and air (39°C, high humidity). Maturation medium consisted of TC199 supplemented with 3ml of bovine LH and FSH (Sioux Biochemical), 0.25mM sodium pyruvate, and 10% fetal calf serum. After 20 hours of incubation in maturation media, oocytes were removed, washed in TL-HEPES and placed in groups of 10 oocytes each into 44 ul microdrops of IVF-Talp (Biowhittaker, Walkersburg, MD) supplemented with 0.22mM sodium pyruvate, and 6 mg/ml essentially fatty acid free BSA. Live sperm were separated by centrifugation in a percoll gradient (Sigma, P1985). Eggs were fertilized by adding one million/ ml of sperm, 2.0ug heparin (Sigma H-3125), and pencillamine, hypotaurine, epinephrine (Sigma-P-4875, H-1384, E-4250) to each drop of fertilization media. Oocytes were removed from fertilization media after 18-22 hours, washed in TL-HEPES, placed in a 1.5 ml tube, and vortexed for 2 min to remove cumulus cells. Twenty-five cumulus free oocytes were placed in each 50ul drop of SOF culture medium, supplemented with 8 mg/ml of fatty acid free BSA, nonessential and essential amino acids, and pyruvate.
Extracted molecule total RNA
Extraction protocol Total RNA extraction was carried out using the mirVana RNA extraction kit according to the manufacturers protocols (Applied Biosciences/Ambion, Austin, TX ).
Label biotin
Label protocol RNA was prepared utilizing three rounds of amplification and hybridized to Affymetrix GeneChipâ Bovine Genome Expression Arrays (Affymetrix, Inc., Santa Clara, CA) utilizing Ambion’s MessageAmp TM II aRNA Amplification and MessageAmpTM II-Biotin Enhanced Kits (Applied Biosystems, Foster City, CA) according to the manufacturers’ instructions.
 
Hybridization protocol Following a 16-hour hybridization the chip was washed and stained utilizing an Affymetrix GeneChipâ Fluidics Station 450 and Affymetrix GeneChipâ Command Console Software. Initial staining utilized streptavidin-conjugated phycoerythrin dye (Invitrogen Corporation, Carlsbad, CA). This stain was enhanced with biotinylated goat anti-streptavidin antibody (Vector Laboratories, Burlingame, CA), then counterstained with the conjugated phycoerythrin.
Scan protocol The chip was scanned with an Affymetrix GeneChipâ Scanner model 3000 7G Plus. Files of the fluorescence intensities were utilized to generate corresponding probe expression levels utilizing Affymetrix Expression Console version 1.1, and quality control values were checked.
Description Gene expression data from oocyte
Data processing The data were analyzed with dChip. Normalization was under default settings.
 
Submission date Sep 23, 2008
Last update date Apr 16, 2009
Contact name Dan Xie
E-mail(s) danxie2@uiuc.edu
Organization name University of Illinois at Urbana and Champaign
Department Bioengineering
Street address 3215 Digital Computer Lab, MC-278, 1304 W. Springfield Ave
City Urbana
State/province IL
ZIP/Postal code 61801
Country USA
 
Platform ID GPL2112
Series (1)
GSE12901 Expression data from early Bovine embryo

Data table header descriptions
ID_REF
VALUE dChip signal intensity

Data table
ID_REF VALUE
AFFX-BioB-5_at 565.67
AFFX-BioB-M_at 825.78
AFFX-BioB-3_at 521.5
AFFX-BioC-5_at 1350.9
AFFX-BioC-3_at 1379.04
AFFX-BioDn-5_at 2625.38
AFFX-BioDn-3_at 3119.08
AFFX-CreX-5_at 11127.29
AFFX-CreX-3_at 11896.41
AFFX-DapX-5_at 1084.57
AFFX-DapX-M_at 1187.21
AFFX-DapX-3_at 4766.88
AFFX-LysX-5_at 57.74
AFFX-LysX-M_at 1224.01
AFFX-LysX-3_at 13077.17
AFFX-PheX-5_at 61.3
AFFX-PheX-M_at 487.97
AFFX-PheX-3_at 8991.55
AFFX-ThrX-5_at 55.15
AFFX-ThrX-M_at 117.18

Total number of rows: 24128

Table truncated, full table size 539 Kbytes.




Supplementary file Size Download File type/resource
GSM323510.CEL.gz 1.9 Mb (ftp)(http) CEL
Processed data included within Sample table

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