|
| Status |
Public on Jun 17, 2014 |
| Title |
28 dpc_crush - left optic nerve |
| Sample type |
RNA |
| |
|
| Source name |
Optic nerves harvested at 28 dpc
|
| Organism |
Mus musculus |
| Characteristics |
strain: BALB/cJ
stress: ONC of left eye
tissue: left optic nerve
timepoint: 28 dpc
|
| Treatment protocol |
The ON of the left eye was crushed 0.5mm posterior from the globe for 4 seconds using the Nickell’s technique. For the microarray studies, five mice were used per time-point. After the ON tissues were harvested samples were all pooled for each experimental (left, n=5) and control eye (right, n=5) group and sent for microarray analysis
|
| Growth protocol |
BALB/cJ mice aged 2-4 months were utilized for all the experiments and were obtained from the Jackson Laboratories.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
ON samples were collected in 1ml of TRIzol and homogenized using 5mm steel beads in the TissueLyser LT for five minutes at 50 oscillations/second. For phase separation 50μl of BAN Phase Separation Reagent was added to the homogenized samples, and samples were centrifuged at 14,000rpm for 15 minutes. The upper aqueous phase was transferred to an RNeasy mini column and processed according to the manufacturer's protocol. Total RNA was pooled from each group (experimental or control group at each time point) and 50ng of RNA from pool was converted to cDNA
|
| Label |
biotin
|
| Label protocol |
Biotinylated cDNA were prepared using NuGen FL-Ovation cDNA Biotin Module according to manufacturers protocol.
|
| |
|
| Hybridization protocol |
Biotin-labeled cDNA was mixed with Affymetrix eukaryotic hybridization buffer, placed onto Affymetrix Mouse Gene 1.0 ST arrays, and incubated at 45º C for 18 h with 60 rpm rotation in an Affymetrix Model 640 Genechip Hybridization Oven. Following hybridization, the arrays were washed, stained with streptavidin-phycoerythrin, and the signal was amplified with an anti-streptavidin antibody using the Affymetrix Model 450 Fluidics Station.
|
| Scan protocol |
Arrays were scanned with the Affymetrix Model 3000 scanner with 7G upgrade
|
| Description |
gene expression data from optic nerve following ONC
|
| Data processing |
Data were collected using the using the GeneChip operating software (GCOS) v1.4 with Affymetrix default analysis settings and global scaling as normalization method
|
| |
|
| Submission date |
Feb 27, 2013 |
| Last update date |
Jun 19, 2014 |
| Contact name |
Tasneem Putliwala Sharma |
| E-mail(s) |
tputliwa@live.unthsc.edu
|
| Phone |
8177352324
|
| Organization name |
UNTHSC
|
| Department |
Cell Biology and Anatomy- Visual Sciences
|
| Lab |
Dr. Abbot F. Clark
|
| Street address |
3500 Camp Bowie Blvd
|
| City |
Fort Worth |
| State/province |
Texas |
| ZIP/Postal code |
76107 |
| Country |
USA |
| |
|
| Platform ID |
GPL6246 |
| Series (2) |
| GSE44705 |
Optic nerve crush induces spatial and temporal gene expression patterns in optic nerve of BALB/cJ mice |
| GSE44708 |
Optic nerve crush induces spatial and temporal gene expression patterns in optic nerve and retina of BALB/cJ mice |
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