|
| Status |
Public on Dec 31, 2013 |
| Title |
Severe-48h-Rep 2 |
| Sample type |
RNA |
| |
|
| Source name |
Rat primary cortical neurons
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Wistar
age: Gestation day 15-16
tissue: Primary cortical neurons
age of culture: Day 21
treatment: Severe
|
| Treatment protocol |
After 21 days in culture, the neurons formed large clusters separated by fasciculated bundles of axons. The axonal bundles were injured by either direct transection with a fine goniotomy knife (Chung et al, 2003, Journal of Neuroscience) or underwent stretch injury via puffing of sterile oxygen (Chung et al, 2005, Journal of Neurotrauma). After the appropriate time, the cells were lysed and mRNA collected using a Qiagen RNeasy isolation kit. At least twelve wells were pooled for each timepoint, and three different cultures from different pregnant rats was used.
|
| Growth protocol |
Cortical neurons were isolated from E17 Hooded Wistar rat embryos as described previously (Chung et al, 2005, Experimental Neurology). Briefly, cortices from embryos were removed and cortical tissue from 8-10 pups was pooled and placed into HEPES buffer with 0.1% trypsin (Sigma) and incubated at 37C for 15 minutes. After three washes with warm HEPES, the tissue was gently triturated and passed through a 20mm gauze filter and plated at a density of 5x105 cells per well onto 19mm2 glass coverslips. Cells were maintained in Neurobasal medium (Gibco) supplemented with 10% B-27 supplement.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted and cleaned up with QIAGEN RNeasy mini kit in accordance with the prescribed protocol provided with the kit. Quality control was performed with E-gene HDA-GT12 genetic analyzer.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays
|
| |
|
| Hybridization protocol |
Standard Illumina hybridization protocol with a hybridization duration of 17h.
|
| Scan protocol |
Standard Illumina scanning protocol with a scan factor = 0.65
|
| Description |
Replicate 2
1901185010_H
|
| Data processing |
The data were normalised using median normalisation with GeneSpring Ver 7.3
|
| |
|
| Submission date |
Aug 18, 2010 |
| Last update date |
Dec 31, 2013 |
| Contact name |
Minghui Jessica Chen |
| Organization name |
Menzies Research Institute
|
| Department |
Neuroscience group
|
| Lab |
A/P Steve Cheung
|
| Street address |
Menzies Research Institute, University of Tasmania, Private Bag 24
|
| City |
Hobart |
| State/province |
Tasmania |
| ZIP/Postal code |
7001 |
| Country |
Australia |
| |
|
| Platform ID |
GPL8031 |
| Series (1) |
| GSE23653 |
Comparative global transcriptomic profiling of mild and severe neurotrauma in an in vitro neuronal model |
|
