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Sample GSM736033 Query DataSets for GSM736033
Status Public on Jun 01, 2014
Title WT, HSP990 treated, rep1
Sample type RNA
 
Source name WT mouse treated with HSP90 inhibitor
Organism Mus musculus
Characteristics strain: WT
treatment: HSP990
gender: male
genetic background: CBA X C57Bl/6
tissue: brain striatum
age: 12 weeks
Treatment protocol HSP990 was obtained from Novartis pharmaceuticals via the CHDI foundation. In order to administer HSP990 to mice, the compound was added to an opaque solution of Saline 10% NVP formulation solution. The mixture was briefly sonicated at high frequency in a water bath and mixed thoroughly to form a uniform suspension. Compound or vehicle alone were administered to mice by oral gavage with thorough mixing performed between dosing to ensure the suspension remained even. Mice were weighed and dosed in the afternoon, and typically dissected at the end of their light cycle.
Growth protocol Mouse housing conditions, genotyping, behavioral assessment, and statistical analysis were performed as per (Hockly et al., 2006; Hockly et al., 2003; Woodman et al., 2007) except that RotaRod and open field analysis were performed over the course of 5 and 30 minutes respectively. Mice were sex matched for each treatment group for all experiments.
Extracted molecule total RNA
Extraction protocol QIAZOL (Qiagen) was added to frozen dissected brain regions (0.2 ml for striatum) prior to homogenisation. The homogenate was flash-frozen and stored at -80oC until all samples are prepared. The homogenate was thawed and placed at room temperature for five minutes to promote dissociation of nucleoprotein complexes. An equal volume of chloroform was added prior to thourough mixing. The homogenate was incubated at room temperature for 3 minutes then centrifuged at 13,000 x g for 15 min at 4°C. The upper aqueous phase was transferred to a new eppendorf and one volume of 70% ethanol was added, then mixed thoroughly. 700μl of sample was transferred to a RNeasy Mini spin column (Qiagen) in a 2ml collection tube. The column/tube assembly was centrifuged for 15 sec at maximum speed at room temperature. The flow-through was discarded and the centrifugation step was repeated with remaining solution from the previous step. 350μl Buffer RW1 was added to the RNeasy spin column and centrifuged for 15 sec at max speed at room temperature prior to discarding the flow-through. DNase treatment was performed using Qiagen RNase-Free DNase Set. For each sample 10μl DNaseI stock solution (lypholised DNase was reconstituted in 550μl RNase free water, aliquoted & stored at -20°C) was diluted in 70μl RDD buffer (i.e. 1:8 dilution), and mixed gently by inverting the tube. 80μl of the DNaseI mix was added to each RNeasy spin column membrane, and incubated at room temperature for 15 min. The washes were performed by adding 350μl Buffer RW1 to the RNeasy spin column prior to centrifuging for 15 sec at max speed at room temperature and discarding the flow-through. 500μl Buffer RPE (with ethanol) was added to the RNeasy spin column, centrifuged for 15 sec at max speed at room temperature and the flow-through was discarded. The step was repeated with a further 500μl Buffer RPE. The RNeasy spin column was placed in new 2ml collection tube & centrifuged for 15 sec at max speed at room temperature to remove any residual ethanol. The RNeasy spin column was placed in a new 1.5ml collection tube and 50μl RNase-free water was added directly to the membrane and incubated at room temperature for 1 min. The column assembly was centrifuged for 15 sec at max speed at room temperature. The elution step was repeated by reusing the eluate. The RNA was stored at -80°C. The RNA was quantified using an Agilent Bioanalyser according to the manufacturer’s instructions prior to cDNA synthesis.
Label biotin
Label protocol Biotinylated cRNAs were prepared from 200ng of total RNA using the GeneChip® 3’ IVT Express Kit (Affymetrix) following the instructions of the manufacturer.
 
Hybridization protocol cRNA (15 ug) was hybridized to GeneChip Mouse Genome 430 2.0 Arrays (Affymetrix) at 45°C for 17 hours, washed and stained according to the protocol described in Affymetrix GeneChip® Expression Analysis Manual (Fluidics protocol FS450_0007).
Scan protocol The arrays were scanned using the GeneChip® Scanner 3000 7G (Affymetrix)
Description Gene expression data
Data processing Microarray quality control was performed using the software package provided on RACE (http://race.unil.ch). Chips with a median normalized unscaled standard error > 1.05 were excluded. Affymetrix annotations (version 30) were used for probeset-to-gene assignments. T-test was performed to assess the differences in gene expression between treated and untreated groups for each genotype.
 
Submission date Jun 01, 2011
Last update date Jun 01, 2014
Contact name Tamara Seredenin
E-mail(s) [email protected]
Organization name EPFL
Street address AI2135 Station 15 EPFL
City Lausanne
ZIP/Postal code 1015
Country Switzerland
 
Platform ID GPL1261
Series (1)
GSE29681 Expression data from WT and R6/2 mice treated with HSP90 inhibitor NVP-HSP990

Data table header descriptions
ID_REF
VALUE RMA signal

Data table
ID_REF VALUE
1415670_at 8.700210857
1415671_at 11.28923777
1415672_at 10.61033591
1415673_at 8.208413741
1415674_a_at 8.707056523
1415675_at 8.713761154
1415676_a_at 10.33451684
1415677_at 8.764966335
1415678_at 10.05618604
1415679_at 10.48586383
1415680_at 7.738394272
1415681_at 9.041402377
1415682_at 8.868790824
1415683_at 9.790766995
1415684_at 7.779297806
1415685_at 8.51298656
1415686_at 9.646546317
1415687_a_at 12.33003304
1415688_at 9.957444927
1415689_s_at 8.57505992

Total number of rows: 45101

Table truncated, full table size 1026 Kbytes.




Supplementary file Size Download File type/resource
GSM736033_WT41.CEL.gz 3.4 Mb (ftp)(http) CEL
Processed data included within Sample table

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