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Sample GSM377758 Query DataSets for GSM377758
Status Public on Mar 05, 2009
Title ReConditioned_Media_rep2_Cy5
Sample type RNA
 
Source name H1 hESC
Organism Homo sapiens
Characteristics cell type: human embryonic stem cells
cell line: H1
Treatment protocol Independent triplicate: groupA in conditioned medium (CM) 9 passages, groupB in 0.2mM sodium butyrate for 6 passages, groupC removed from 4 passages in butyrate back into CM for 3 more passages
Growth protocol hESC in DMEM/F12, 20% serum replacer, glutamax, pen/strep, NEAA, pyruvate (Invitrogen) and mercaptoethanol (Sigma), grown on Matrigel (BD) 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 in Butyrate followed by reconditioned media
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 GPL1708
Series (2)
GSE15110 H1 hESC Exposed to Butyrate
GSE15112 ESC Exposed to Butyrate

Data table header descriptions
ID_REF
VALUE Normalized signal intensity

Data table
ID_REF VALUE
3 8.99
4 8.962
5 8.983
6 8.914
8 6.157
9 8.147
10 9.685
12 5.691
13 6.288
15 5.704
16 5.128
17 8.659
18 10.147
19 11.995
20 10.955
22 5.745
23 11.114
24 11.192
25 13.802
26 9.428

Total number of rows: 41675

Table truncated, full table size 490 Kbytes.




Supplementary file Size Download File type/resource
GSM377758_US23502338_251239137853_S01_A01_Cy5.txt.gz 10.9 Mb (ftp)(http) TXT
Processed data included within Sample table

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