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Sample GSM377740 Query DataSets for GSM377740
Status Public on Mar 05, 2009
Title Butyrate_rep2_Cy5
Sample type RNA
 
Source name R1 mESC
Organism Mus musculus
Characteristics cell type: mouse embryo stem cells
cell line: R1
Treatment protocol Independent quadruplicate: groupA in LIF (Chemicon) without feeders, groupB in 0.2mM sodium butyrate (without LIF) for 15 passages
Growth protocol mESC in DMEM, 15% serum replacer, glutamax, pen/strep, NEAA, pyruvate (Invitrogen) and mercaptoethanol (Sigma), grown on gelatin (Sigma) at 37C in 5% CO2
Extracted molecule total RNA
Extraction protocol RNA extraction utilized the Rneasy Minikit (Qiagen) following manufacturer's protocol. The RNA was treated with Dnase and given to the Center for Array Technologies for quantitation and purity analysis.
Label Cy5
Label protocol Cyanine-3 (Cy3) and Cyanine-5 (Cy5) labeled cRNA was prepared from 0.5 ug RNA using the Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
 
Hybridization protocol 825ng of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) and 825ng of Cy5-labelled cRNA (specific activity >10.0 pmol Cy5/ug cRNA) were combined and fragmented at 60°C for 30 minutes in a reaction volume of 55 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) and Agilent Whole Genome Mouse Microarrays (G4122F) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by compressed nitrogen (N2) gun.
Scan protocol Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using two color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Red and both Green PMT and Red PMT arer set to 100%).
Description Gene Expression after exposure to Butyrate
Data processing The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters (protocol GE2-v5_95 and Grid: 014868_D_20060807) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
 
Submission date Mar 04, 2009
Last update date Mar 04, 2009
Contact name Brig Mecham
E-mail(s) bmecham@fhcrc.org
Organization name University of Washington
Street address 1100 Fairview Ave N
City Seattle
State/province WA
ZIP/Postal code 98109
Country USA
 
Platform ID GPL4134
Series (2)
GSE15107 R1 mESC Exposed to Butyrate
GSE15112 ESC Exposed to Butyrate

Data table header descriptions
ID_REF
VALUE Normalized signal intensity

Data table
ID_REF VALUE
12 2.693
13 6.187
14 2.745
15 2.714
16 10.691
18 2.735
19 2.741
20 2.747
21 2.753
22 10.968
23 2.764
24 8.602
25 5.509
26 9.029
27 8.615
28 4.619
29 8.972
30 2.794
31 2.798
32 2.802

Total number of rows: 43379

Table truncated, full table size 504 Kbytes.




Supplementary file Size Download File type/resource
GSM377740_US23502338_251486811295_S01_H_GE2-v5_95_Feb07_1_2_Cy5.txt.gz 11.3 Mb (ftp)(http) TXT
Processed data included within Sample table

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