|
| Status |
Public on Aug 31, 2007 |
| Title |
Mouse young brain C5-Br-3 |
| Sample type |
RNA |
| |
|
| Source name |
Young neocortex
|
| Organism |
Mus musculus |
| Characteristics |
strain: B6/C3H F1 hybrid
gender: male
age: 5-month-old
|
| Biomaterial provider |
University of Wisconsin-Madison, Genetics
|
| Treatment protocol |
control diet
|
| Growth protocol |
Male B6C3F1 mice were purchased from Harlan Sprague-Dawley at 6-7 weeks of age. Mice were housed singly in a pathogen-free facility and provided acidified water ad libitum.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was extracted from frozen tissue using the TRIZOL reagent (Life Technologies, Grand Island, NY) and mRNA was purified from the total RNA with oligo-dT-linked oligotex resin (Qiagen, Valencia, CA). Then, double-stranded cDNA was synthesized from 1ug of mRNA using the SuperScript Choice System (Life Technologies, Grand Island, NY) with an oligo-dT primer containing a T7 RNA polymerase promoter (Genset, La Jolla, CA), purified with phenol-chloroform-isoamyl alcohol, and precipitated with pellet paint co-precipitant (Novagen, Madison, WI). Purified double-stranded cDNA was used to synthesize biotin-labeled cRNA using the BioArray High Yield RNA Transcript Labeling Kit (Enzo, Farmingdale, NY). The biotin-labeled antisense cRNA was then purified with the RNeasy affinity column (Qiagen, Valencia, CA). After purification, biotin-labeled cRNA was fragmented randomly to sizes ranging from 35 to 200 bases.
|
| Label |
C5-Br-3
|
| Label protocol |
C: control diet
5: 5-month-old
Br: brain(neocortex)
3: sample number
|
| |
|
| Hybridization protocol |
Total 10ug cRNA was placed in the gene chip. The gene chip I used for this study is the Affymetrix MOE 430 2.0 arrays (Affymetrix, Santa Clara, CA). The Affymetrix MOE 430 2.0 array has 45,037 probe sets. Hybridization was carried out at 45ºC for 16 h with mixing on a rotisserie at 60 rpm. Following hybridization, the hybridization solutions were removed and the gene chips were be installed in a fluidics system for washes and staining.
|
| Scan protocol |
After staining, the gene chips were read at a resolution of 3 μm using a Hewlett Packard GeneArray Scanner (Affymetrix, Santa Clara, CA). The averaged images collected from two scanned images were used for the analysis.
|
| Description |
We used five animals per group, and hybridized each sample to independent DNA chips, because previous work from our laboratory suggests that variability between individuals is higher than variability observed in replicate hybridizations of the same samples.
|
| Data processing |
The GeneChip Operating Software (GCOS) version 1.2 was used to analyze the image data. GCOS determines the presence of mRNA in samples and computes the signals of probe sets using the differences and ratios between PM and MM signals. Signals in each image are normalized to minimize an overall variability in hybridization intensities by a global scaling method.
|
| |
|
| Submission date |
Jun 15, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Sang-Kyu Park |
| E-mail(s) |
sangkyu@colorado.edu
|
| Phone |
303-492-5159
|
| Fax |
303-492-8063
|
| Organization name |
University of Colorado at Boulder
|
| Department |
IBG
|
| Street address |
1480 30th St.
|
| City |
Boulder |
| State/province |
CO |
| ZIP/Postal code |
80303 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (2) |
| GSE8150 |
Age-related transcriptional changes and the effect of dietary supplementation of vitamin E in the mouse brain |
| GSE8162 |
Age-related transcriptional changes and the effect of dietary supplementation of vitamin E in the mouse heart and brain |
|
