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Sample GSM1151218 Query DataSets for GSM1151218
Status Public on May 31, 2013
Title WT8S
Sample type RNA
Source name primary mouse embryonic fibroblasts (MEFs) from E10.0 embryo obtained from Rb-ATM double knockout C57BL/6J mouse
Organism Mus musculus
Characteristics background strain: C57BL/6J
cell type: primary mouse embryonic fibroblasts (MEFs)
transfection: vector expressing ATM
genotype/variation: Rb-/-;ATM-/-;ATM-reconstituted
Treatment protocol Primary MEFs were transduced with empty vector or vector expressing ATM or vector expressing DNMT1 shRNA, selected with 4μg/ml puromycin for 3 days, grown under normal culture conditions for another 4 days and analysed.
Growth protocol Primary MEFs were grown in MEM-α supplemented with 10% FBS under normal culture conditions.
Extracted molecule total RNA
Extraction protocol RNA isolation RNA was quantified and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
Label Cy3
Label protocol Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng of total RNA using the One-Color Low Input Quick Amp Labeling kit (Agilent) according to the manufacturer's instructions with some changes. 3rd Spike mix was made by diluted 2nd Spike-Mix 10 times, and 1.8 ul of 3rd Spike-Mix was added to samples. Cy3 labeled cRNA was synthesized at 40°C for 20 hours and purified by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
Hybridization protocol 1.65 ug of Cy3-labelled cRNA (specific activity >7.0 pmol Cy3/ug cRNA) was 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 Mouse Whole Genome Microarray 4x44K (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 by raise it slowly from buffer 2.
Scan protocol Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5um, Dye channel is set to Green. Scan mode is eXtended Dynamic range Scan mode and PMT is set to 10% and 100%).
Description 163 cells reconstituted with vector expressing ATM, selected with 4μg/ml puromycin for 3 days, grown under normal culture conditions for another 4 days and analysed.
Data processing The scanned images were analyzed with Feature Extraction Software (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 014868_D_F_20100617) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
Submission date May 30, 2013
Last update date May 31, 2013
Contact name Awad Shamma
Phone +81-76-264-6751
Fax +81-76-234-4521
Organization name Kanazawa Uiniversity
Department Cancer Research Institute
Lab Division of Oncology and Molecular Biology
Street address Kakuma-machi
City Kanazawa
State/province Ishikawa
ZIP/Postal code 920-1192
Country Japan
Platform ID GPL10333
Series (1)
GSE47501 ATM mediates pRB function to control DNMT1 protein stability and DNA methylation

Data table header descriptions
VALUE Normalized Signal intensity

Data table
1 1.81E+04
2 8.12E+00
3 8.16E+00
4 8.20E+00
5 8.23E+00
6 8.27E+00
7 8.30E+00
8 8.33E+00
9 8.36E+00
10 8.38E+00
11 8.41E+00
12 8.45E+00
13 8.45E+00
14 8.47E+00
15 1.80E+01
16 7.37E+03
17 1.12E+05
18 8.55E+00
19 8.56E+00
20 1.03E+05

Total number of rows: 44397

Table truncated, full table size 639 Kbytes.

Supplementary file Size Download File type/resource
GSM1151218_US22502564_252665510305_S01_GE1_107_Sep09_1_1.txt.gz 2.2 Mb (ftp)(http) TXT
Processed data included within Sample table

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