|
| Status |
Public on Jun 21, 2014 |
| Title |
Intact,SN,Rep3 |
| Sample type |
RNA |
| |
|
| Source name |
adult male Sprague Dawley rat
|
| Organism |
Rattus norvegicus |
| Characteristics |
tissue: 1mm3 tissue punch from left side substantia nigra
gender: male
developmental stage: adult
strain: Sprague-Dawley
|
| Treatment protocol |
Rats were anesthetized prior to surgery with Equi-Thesin (0.3 ml/100 g body weight i.p.; chloral hydrate 42.5 mg/ml + sodium pentobarbital 9.72 mg/ml) and rats injected in two sites in the striatum with either 6OHDA (MP Biomedicals, Solon, OH; 5 μg/μl 6-OHDA in 0.02% ascorbic acid, 0.9% saline solution) or vehicle. The coordinates for these injections were AP +1.6 mm, ML +2.4 mm, DV −4.2 mm and AP −0.2 mm, ML + 2.6 mm, DV −7.0 mm. A total of 2 μl 6-OHDA (total dose of 6-OHDA = 20 μg over two sites) or vehicle was injected at 0.5 μl/minute.
|
| Growth protocol |
Male, Sprague-Dawley rats weighing between 200–225 g were given access to food and water ad libitum, and housed in an AAALAC accredited animal facility on a standard light-dark cycle. Housing conditions and animal procedures were performed according to AAALAC guidelines.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Rats were sac’d, brains frozen in isopentane for 30 seconds and stored individually at -80˚C. Substantia Nigras were dissected using punches of 1 cubic mm at -20˚C in a cryostat. Punches were ejected into an eppendorf tube containing Trizol and gently homogenized using a disposable plastic pestle. Samples were stored at -80˚C. RNA was isolated from trizol homogenate using RNA Clean and Concentrator kit (Zymo). Quality of total RNA was determined on an Agilent Bioanalyzer.
|
| Label |
biotin
|
| Label protocol |
Biotin labeled, cRNA was made using Ambion WT Expression kit (Life Technologies) combined with the GeneChip WT Terminal Labeling Kit (Affymetrix).
|
| |
|
| Hybridization protocol |
2.5 ug of fragmented cRNA was hybridized for 17 hours at 45C in the GeneChip Hybridization Oven 640 (Affymetrix). Chips were washed and stained in the Fluidics Station 450 (Affymetrix) using fluidix protocol FS450-0007.
|
| Scan protocol |
Genechips were scanned in a Gene Chip Scanner 3000 7G (Affymetrix).
|
| Data processing |
Only samples meeting all of the quality control measures defined by Affymetrics were included in this study. Multidimensional plots and Pearson Correlations were used to establish group/treatment homogeneity.
|
| |
|
| Submission date |
Jun 20, 2014 |
| Last update date |
Jun 21, 2014 |
| Contact name |
Jack W Lipton |
| E-mail(s) |
[email protected]
|
| Phone |
616-234-0950
|
| Organization name |
Michigan State University
|
| Department |
TSMM
|
| Street address |
333 Bostwick Ave.
|
| City |
Grand Rapids |
| State/province |
MI |
| ZIP/Postal code |
49503 |
| Country |
USA |
| |
|
| Platform ID |
GPL6247 |
| Series (1) |
| GSE58710 |
Time Course of Gene Expression in the Substantia Nigra in Response to Intrastriatal 6-hydroxydopamine in the rat. |
|
