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Sample GSM979003 Query DataSets for GSM979003
Status Public on Aug 03, 2012
Title 2hr flagellin exposure replicate1 vs 0hr untreatedpool
Sample type RNA
 
Channel 1
Source name RAW 264.7 cell line
Organism Mus musculus
Characteristics cell line: RAW 264.7
treatment: 2hr flagellin exposure
Treatment protocol Cells were treated with either flagellin or LPS for 1, 2, 4, or 24 hours.
Growth protocol RAW 264.7 cells were grown in DMEM supplemented with 10% FBS.
Extracted molecule total RNA
Extraction protocol Cells were washed twice with PBS and then RNA was harvested according to the TriZol protocol.
Label Cy5
Label protocol A total of 2 ug of RNA in a mixture containing 6 ug of random hexamers, 0.01 M dithiothreitol, an aminoallyl-deoxynucleoside triphosphate mixture containing 25 mM each dATP, dCTP, and dGTP, 15 mM dTTP, and 10 mM amini-allyl dUTP (aa-dUTP), reaction buffer, and 400 units of SuperScript III reverse transcriptase at 42C overnight. The RNA template then was hydrolyzed by adding NaOH and EDTA to a final concentration of 0.2 and 0.1 M, respectively, and incubating at 65C for 15 min. Unincorporated aa-UTP was removed with a MinElute column. The probe was eluted with a phosphate elution buffer (4 mM KPO4, pH 8.5, in ultrapure water), dried and resuspended in 0.1 M sodium carbonate buffer (ph 9.0). To couple the amino-allyl cDNA with fluorescent labels, normal human serum-Cy3 or normal human serum-Cy5 was added at room temperature for 1 hour. Uncoupled label was removed using the Qiagen MinElute columns.
 
Channel 2
Source name RAW 264.7 cell line
Organism Mus musculus
Characteristics cell line: RAW 264.7
treatment: none
Treatment protocol Cells were treated with either flagellin or LPS for 1, 2, 4, or 24 hours.
Growth protocol RAW 264.7 cells were grown in DMEM supplemented with 10% FBS.
Extracted molecule total RNA
Extraction protocol Cells were washed twice with PBS and then RNA was harvested according to the TriZol protocol.
Label Cy3
Label protocol A total of 2 ug of RNA in a mixture containing 6 ug of random hexamers, 0.01 M dithiothreitol, an aminoallyl-deoxynucleoside triphosphate mixture containing 25 mM each dATP, dCTP, and dGTP, 15 mM dTTP, and 10 mM amini-allyl dUTP (aa-dUTP), reaction buffer, and 400 units of SuperScript III reverse transcriptase at 42C overnight. The RNA template then was hydrolyzed by adding NaOH and EDTA to a final concentration of 0.2 and 0.1 M, respectively, and incubating at 65C for 15 min. Unincorporated aa-UTP was removed with a MinElute column. The probe was eluted with a phosphate elution buffer (4 mM KPO4, pH 8.5, in ultrapure water), dried and resuspended in 0.1 M sodium carbonate buffer (ph 9.0). To couple the amino-allyl cDNA with fluorescent labels, normal human serum-Cy3 or normal human serum-Cy5 was added at room temperature for 1 hour. Uncoupled label was removed using the Qiagen MinElute columns.
 
 
Hybridization protocol Labeled cDNA was hybridized to the Agilent arrays according to the Agilent protocol.
Scan protocol Hybridizations were scanned on the Axon 4000B scanner with GenePix Pro software.
Data processing Intensity values were extracted using Feature Extraction 9.5.3.1 from Agilent and then normalized and analyzed using MIDAS and MeV from the TM4 Suite.
 
Submission date Jul 31, 2012
Last update date Aug 03, 2012
Contact name Bryan Frank
E-mail(s) bfrank@jcvi.org
Phone 301-795-7000
Organization name The J. Craig Venter Institute
Department Dept of Infectious Diseases
Street address 9704 Medical Center Drive
City Rockville
State/province MD
ZIP/Postal code 20850
Country USA
 
Platform ID GPL10333
Series (1)
GSE39785 Model-driven multi-omic data analysis elucidates metabolic immunomodulators of macrophage activation

Data table header descriptions
ID_REF
VALUE log2 normalized ratio of test (Cy5,IB)/reference (Cy3,IA)
PRE_VALUE test (IB channel) / reference (IA channel)

Data table
ID_REF VALUE PRE_VALUE
1 0.0393 1.027639504
2 0.0186 1.012957317
3 -0.1014 0.932105869
4 -0.2993 0.812627291
5 0.0166 1.011573237
6 -0.0465 0.968294461
7 -0.1481 0.902420575
8 -0.0669 0.954697987
9 0.1271 1.092109961
10 -0.1076 0.928121895
11 -0.1750 0.885775098
12 -0.0386 0.97363266
13 0.4122 1.330713329
14 0.5588 1.473017697
15 0.2407 1.181595692
16 -0.1221 0.91887209
17 -0.4447 0.73475092
18 0.1089 1.078381795
19 0.4674 1.382646913
20 -0.0303 0.979220251

Total number of rows: 44397

Table truncated, full table size 1092 Kbytes.




Supplementary file Size Download File type/resource
GSM979003_2hr_flagellin_R1_vs_0hr_untreatedpool_252665510510_1_2.mev.gz 1.3 Mb (ftp)(http) MEV
GSM979003_2hr_flagellin_R1_vs_0hr_untreatedpool_252665510510_1_2.txt.gz 15.8 Mb (ftp)(http) TXT
Processed data included within Sample table
Processed data provided as supplementary file

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