|
| Status |
Public on Nov 21, 2011 |
| Title |
TES 3 4h |
| Sample type |
RNA |
| |
|
| Source name |
retina
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Brown Norway Rat
gender: male
age: 8 weeks
|
| Extracted molecule |
total RNA |
| Extraction protocol |
mRNA of retina was isolated 4h after transcornial electrical stimulation and compared to sham animals using RNAeasy Isolation Kit (Ambion, Austin, Texas, USA) according to manufacturers instructions.
|
| Label |
Biotin
|
| Label protocol |
Three microliter total RNA was used for each sample to obtain linearly amplified labeled cRNA by using GeneChip® One-Cycle Target Labeling kit (Affymetrix, Santa Clara, CA, USA) as described by manufacturer. Briefly, total RNA was used to generate double-stranded cDNA with the T7-oligo (dT) primer. This double-stranded cDNA was used in vitro transcription and biotin labeling steps. Labeled cRNA yield and purity were determined by measuring the absorbance at 260 and 280nm. All the cRNA 260/280 ratios were between 1.9 and 2.1. Quality control of the labeled cRNA products was assessed by performing 1µg labeled cRNA on 2% agarose gel to see similar RNA smear type.
|
| |
|
| Hybridization protocol |
15 micrograms of labeled cRNA from retina was fragmented, and 10ug was hybridized to Affymetrix Rat Gene 1.0 ST Gene Chip arrays for 18-24h (Affymetrix Inc.). Gene chip was washed and stained with streptavidin-phycoerythrin.
|
| Scan protocol |
The chip was scanned at 6um resolution with Agilant model G2500A Gene Array scanner. A visual quality control measurement was performed to ensure proper hybridization after each chip was scanned.
|
| Data processing |
For statistical analysis of microarray data all .cel files were imported to GeneSpring GX software (Agilent Technologies Inc.) where GC-RMA algorithm was applied to yield log signal values on each probe set. All transcripts with a minimum change of 1.2 fold together with a p value less than 0.05 were selected using Significance Analysis of Microarray (SAM) (Tusher VG, 2001).
|
| |
|
| Submission date |
Dec 20, 2010 |
| Last update date |
Nov 21, 2011 |
| Contact name |
Gabriel Willmann |
| E-mail(s) |
[email protected]
|
| Organization name |
University Eye Hospital and Centre for Ophthalmology Tübingen
|
| Street address |
Schleichstrasse 12-16
|
| City |
Tübingen |
| ZIP/Postal code |
72076 |
| Country |
Germany |
| |
|
| Platform ID |
GPL10741 |
| Series (1) |
| GSE26174 |
Gene Expression Profiling of the Retina after Transcorneal Electrical Stimulation in Wildtype Brown Norway Rats |
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