|
| Status |
Public on May 12, 2015 |
| Title |
DRG_RTX_100nM_Rep4 |
| Sample type |
RNA |
| |
|
| Source name |
Dissociated dorsal root ganglia treated for 30 min with RTX
|
| Organism |
Rattus norvegicus |
| Characteristics |
gender: male
age: 8 weeks
strain: Sprague Dawley rats
tissue: lumbar dorsal root ganglia
|
| Treatment protocol |
Dorsal root ganglia were incubated in 5 ml MEM containing collagenase P (Roche, Penzberg, DE) (0.1 U/ml, 1 h, 37°C, 5% CO2), dissociated by trituration with fire-polished Pasteur pipettes, and then treated with the solvent DMSO (0.1%) or RTX (100 nM). The cells were spun down (100 g, 5 min), resuspended in 1 ml Hanks buffered salt solution (pH 8.0) containing 0.025% EDTA and 245 U/ml trypsin (Worthington, Lakewood, NJ), and incubated for 4 min at 37°C in a water bath. Trypsination was stopped by the addition MgSO4 (400 µM final conc.). The cell suspension was then loaded onto BSA gradients (15% BSA in Hanks buffered salt solution) and centrifuged (120 g, 8 min). The pellet was frozen in liquid nitrogen and stored at -80°C for RNA isolation.
|
| Growth protocol |
Lumbar dorsal root ganglia were isolated from 8 weeks old Spargue Dawley rats
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA isolation was performed with NucleoSpin RNA/Protein kits from Macherey-Nagel including on column DNase treatment. The RNA concentration and quality was measured with spectroscopy (Nanodrop, Thermo Fisher Scientific) and capillary electrophoresis (Agilent 2100 Bioanalyzer, 28S:18S rRNA ratio > 1.7, RNA integrity number (RIN) > 8.2).
|
| Label |
Cy3
|
| Label protocol |
Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays
|
| |
|
| Hybridization protocol |
standard as recommended by illumina
|
| Scan protocol |
standard as recommended by illumina
|
| Description |
Method to eliminate TRPV1-positive neurons
|
| Data processing |
Raw fluorescence intensities were background corrected and exported from the Illumina Genome studio software. Background corrected data were log2 transformed and normalized (quantile method) using the R package lummi. The data were filtered for expressed genes (detection p < 0.01) and differentially expressed transcripts were identified with the R package limma. P-values were adjusted for multiple testing with Benjamini and Hochberg's method.
|
| |
|
| Submission date |
Jul 23, 2014 |
| Last update date |
May 12, 2015 |
| Contact name |
Joerg Isensee |
| E-mail(s) |
joerg.isensee@uk-koeln.de
|
| Phone |
+4922147887591
|
| Organization name |
University Hospital of Cologne
|
| Department |
Department of Anesthesiology and Intensive Care Medicine
|
| Lab |
Experimental Anesthesiology and Pain Research
|
| Street address |
Robert Koch Str. 10
|
| City |
Cologne |
| ZIP/Postal code |
50931 |
| Country |
Germany |
| |
|
| Platform ID |
GPL6101 |
| Series (1) |
| GSE59727 |
The transcriptome of TRPV1-positive sensory neurons revealed by subgroup-elimination transcriptomics |
|
