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Sample GSM154523 Query DataSets for GSM154523
Status Public on Jan 16, 2007
Title Serum starved MEF, newly transcribed RNA, biological rep2
Sample type RNA
Source name 48h serum starved primary mouse embryonic fibroblasts (MEF)
Organism Mus musculus
Characteristics Genotype: C57/BLACK6, ATF1 -/+
Age: e14.5
Treatment protocol Primary mouse embryonic fibroblasts were isolated as described previously (Todaro G.J. and Green H. (1963), J. Cell Biol., 17, 299-313; Brusselbach S.U. et al. (1995), Oncogene, 10, 79-86.).
Growth protocol Cells were grown using high glucose-DMEM (Invitrogen) with 1% Penicillin/Streptomycin (Invitrogen) and 10% dialyzed FBS (cutoff 10,000Da, Sigma) to near confluency. Cells were serum-starved for 48h with 1% dialyzed FBS, and then incubated for 2h with media containing 200µM 4-thiouridine (s4U, Sigma), 1% FBS and 1µCi 3H-cytidine (20Ci/mmol, Biotrend).
Extracted molecule total RNA
Extraction protocol RNA was isolated according to the Qiagen RNeasy Midi protocol. RNA quality and yield were measured by the RNA6000 Nano assay (Bioanalyzer 2100, Agilent Technologies). 100µg of RNA were diluted in binding buffer and loaded onto ~150µl packaged volume of the equilibrated organomercurial matrix (Squarix Biotechnology Germany) loaded to a Small Spin Column (Qiagen). After 4h incubation under constant gentle rotation and light protection at 4°C, non-bound RNA was eliminated by sequential washing steps. Bound (thiolated) RNA was eluted using binding buffer supplemented with 20mM 2-mercaptoethanol and ethanol precipitated. Specific activity of eluted RNA was calculated via 3H-cytidine measurement on a Beckman Coulter LS6500.
Label biotin
Label protocol RNA was reverse-transcribed, linearly amplified as described previously (Kenzelmann M. et al. (2004), Genomics 83, 550-558.), and biotin-labelled via a T7-based strategy according to manufacturer's instructions (Affymetrix).
Hybridization protocol Following fragmentation, 10µg of cRNA were hybridized to MU74Av2 GeneChip (Affymetrix) according to manufacturer's instructions.
Scan protocol GeneChips were scanned using the GeneChip' Scanner 3000 7G System (Affymetrix).
Description Gene expression data from primary serum starved MEF cells grown to near confluency.
Data processing The data were analyzed with the SAS MicroArray Solution 1.0, using standard settings except the following: consistence within experimental groups (stimulus and labelling combination) was determined by array group correlation. Loglinear mixed models with Bonferroni corrections were fitted for values of perfect-matches, only with stimulus (10% FBS) and labelling (s4U) considered to be constant and the chip-ID as random.
Submission date Jan 10, 2007
Last update date Jan 10, 2007
Contact name Marc Kenzelmann
Phone +41 31 632 3251
Fax +41 31 632 3550
Organization name Institute of Medical Microbiology, University of Bern
Department Molecular Genome Analysis
Street address Friedbuhlstrasse 51
City Bern
ZIP/Postal code 3010
Country Switzerland
Platform ID GPL81
Series (1)
GSE6697 Expression data from spec. transcriptional activity of primary mouse embryonic fibroblasts (MEF) in response to serum.

Data table header descriptions
VALUE MAS5-calculated Signal intensity
ABS_CALL the call in an absolute analysis that indicates if the transcript was present (P), absent (A), marginal (M), or no call (NC)
DETECTION P-VALUE 'detection p-value', p-value that indicates the significance level of the detection call

Data table
AFFX-MurIL2_at 2.1 A 0.860518
AFFX-MurIL10_at 1.6 A 0.876428
AFFX-MurIL4_at 3 A 0.645547
AFFX-MurFAS_at 317.8 P 0.00039
AFFX-BioB-5_at 104.9 P 0.001102
AFFX-BioB-M_at 240.7 P 4.4e-05
AFFX-BioB-3_at 123.4 P 0.00011
AFFX-BioC-5_at 437.1 P 7e-05
AFFX-BioC-3_at 353.5 P 5.2e-05
AFFX-BioDn-5_at 648.5 P 4.4e-05
AFFX-BioDn-3_at 1926.9 P 4.4e-05
AFFX-CreX-5_at 3552.5 P 4.4e-05
AFFX-CreX-3_at 5899.3 P 4.4e-05
AFFX-BioB-5_st 9.3 A 0.227636
AFFX-BioB-M_st 3 A 0.659339
AFFX-BioB-3_st 4 A 0.794268
AFFX-BioC-5_st 2.4 A 0.852061
AFFX-BioC-3_st 1.3 A 0.904333
AFFX-BioDn-5_st 3.4 A 0.51489
AFFX-BioDn-3_st 3.5 A 0.749204

Total number of rows: 12488

Table truncated, full table size 315 Kbytes.

Supplementary file Size Download File type/resource
GSM154523.cel.gz 2.8 Mb (ftp)(http) CEL

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