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Sample GSM154522 Query DataSets for GSM154522
Status Public on Jan 16, 2007
Title Serum treated MEF, newly transcribed RNA, biological rep1
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), 10% 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 treated 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
E-mail(s) m.kenzelmann@dkfz.de
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
ID_REF
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
ID_REF VALUE ABS_CALL DETECTION P-VALUE
AFFX-MurIL2_at 1.9 A 0.860518
AFFX-MurIL10_at 2.7 A 0.645547
AFFX-MurIL4_at 0.8 A 0.916408
AFFX-MurFAS_at 35.2 P 0.020022
AFFX-BioB-5_at 150.3 P 0.00039
AFFX-BioB-M_at 354.6 P 4.4e-05
AFFX-BioB-3_at 274.3 P 4.4e-05
AFFX-BioC-5_at 632.9 P 4.4e-05
AFFX-BioC-3_at 371.5 P 4.4e-05
AFFX-BioDn-5_at 939.1 P 4.4e-05
AFFX-BioDn-3_at 1962.7 P 4.4e-05
AFFX-CreX-5_at 5130.8 P 4.4e-05
AFFX-CreX-3_at 6040.3 P 4.4e-05
AFFX-BioB-5_st 16.7 P 0.026111
AFFX-BioB-M_st 2.8 A 0.749204
AFFX-BioB-3_st 4.8 A 0.794268
AFFX-BioC-5_st 3.8 A 0.868639
AFFX-BioC-3_st 0.8 A 0.966323
AFFX-BioDn-5_st 3.1 A 0.631562
AFFX-BioDn-3_st 5.5 A 0.544587

Total number of rows: 12488

Table truncated, full table size 312 Kbytes.




Supplementary file Size Download File type/resource
GSM154522.cel.gz 2.7 Mb (ftp)(http) CEL

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