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Sample GSM586849 Query DataSets for GSM586849
Status Public on Nov 17, 2011
Title C2C12_cells_treated_with_rhBMP2
Sample type RNA
 
Source name C2C12 cells treated with rhBMP2
Organism Mus musculus
Characteristics cell type: pre-myoblastic and pre-osteoblastic C2C12 cell line
Biomaterial provider American Type Culture Collection
Treatment protocol C2C12 cultures treated for 12h with 200ng/ml rhBMP2
Extracted molecule total RNA
Extraction protocol total RNA was extracted with RNeasy
The cRNA was prepared as recommended in the Amersham CodelinkTM iExpress Assay Reagent Kit manual. Briefly, the sample containing 1.5 µg RNA of culture treated for 12h with rhBMP2 was mixed with 7.5 µl of
bacterial mRNAs (10pg/µl), 1 µl of oligo dT in a final volume of 12 µl. The reaction was incubated at 70ºC for 10min and kept on ice by 3min. Subsequently, 2 µl of 10X First Strand Buffer, 4 µl 5mM dNTPs, 1 µl RNase inhibitor, and 1 µl ArrayScript were added to tube, in a final volume of 20 µl. The reaction was incubated at 42°C for 2h. After this period, the tube was placed on ice and 63 µl water, 10 µl of 10X Second Strand Buffer, 4 µl of 5mM dNTPs, 2 µl of DNA polymerase and 1 µl of RNaseH were added. The reaction is incubating at 16°C for 2h.
Label biotin
Label protocol The cDNA was purified and 20 µl of each purified cDNA was mixed with 12 µl Biotin-
NTP, 4µl of 10X T7 reaction buffer and 4 µl of 10X Enzyme Mix reaction buffer and incubated at 37°C for 14h. After cRNA synthesis, the cRNA preparation was purified
and 10 µg of biotinylated cRNA was mixed with 5 µl of Fragmentation Buffer in a final volume of 25 µl , incubated at 94°C for 20min and then kept on ice for 5min.
 
Hybridization protocol 25µl of reaction was transferred to another tube containing 78µl of Hybridization Buffer A, 130µl of Hybridization Buffer B and 27µl of water and the reaction was incubated at 90°C for 5min. Each tube was then placed on ice and then applied to the Codelink DNA microarray containing around 36,000 genes. The subsequent stages of hybridization, washing and analysis of each microarray were carried out as detailed in the Codelink Gene Expression System: Single-Assay Biorray Hybridization and Detection manual.
Scan protocol Scanner Gene Pix 4000B (Molecular Devices) standard procedure
Description n/a
Data processing The gene expression date were normalized using the Suport Vector Regression (SVR). Implemented in the GEDI toolbox (http://www.iq.usp.br/wwwdocentes/mcsoga/gedi/) . Identification of these genes was obtained using the ratio of normalized intensities between rhBMP2 and control (untreated) samples, considering a fold change of >= 2.5.
 
Submission date Aug 24, 2010
Last update date Nov 17, 2011
Contact name Maria Angélica Lopes de Souza
Organization name Scylla Bioinformática
Street address Franscisco Otaviano nº 60 sl 52
City Campinas
State/province São Paulo
ZIP/Postal code 13083765
Country Brazil
 
Platform ID GPL8063
Series (1)
GSE23783 Genes modulated by rhBMP2 and rhBMP7 during osteoblastic differentiation of C2C12 cells

Data table header descriptions
ID_REF
SIGNAL_RAW Raw Intensity
VALUE Normalized with SVR

Data table
ID_REF SIGNAL_RAW VALUE
1 19.97 20.8616037
2 4215.76 3925.821773
3 121.41 112.6392107
4 32.49 32.55347265
5 91.35 85.71699703
6 1216.59 1097.575989
7 45.3 44.64595167
8 1548.86 1401.630006
9 9021.02 8566.873396
10 1744.6 1583.529474
11 2104.8 1924.852617
12 24.8 26.01302287
13 1518.34 1374.858972
14 1143.56 1030.902371
15 705.74 633.0431529
16 256.07 231.9725577
17 3605.49 3339.336549
18 1143.84 1031.260764
19 633.17 567.8389544
20 13.06 14.16256632

Total number of rows: 36227

Table truncated, full table size 836 Kbytes.




Supplementary file Size Download File type/resource
GSM586849.txt.gz 1.6 Mb (ftp)(http) TXT
Processed data included within Sample table

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