|
| Status |
Public on Jan 31, 2016 |
| Title |
T_240649 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
TRP allograft tumor_untreated
|
| Organism |
Mus musculus |
| Characteristics |
strain background: C57BL/6
mouse id: 240649
survival (days): 25
treatment: untreated
tissue: high-grade astrocytoma allografts
|
| Treatment protocol |
Tumors were harvested from bains of mice and immediately flash-frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted s with Qiagen RNeasy kit according to manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
Labeling was performed according to the Low RNA Input Linear Amplification kit from Agilent. 1µg of RNA was primed with 1.2 µl of provided T7 promoter primer at 65 °C for 10 minutes. Reverse transcription was performed with provided Moloney Murine Leukemia Virus Reverse Transcriptase at 40 °C for 2 hours, 65 °C for 15 minutes, and 4 °C for 5 minutes. Laeled RNA was made by adding 1µl of 10mM Cy3 or Cy5 added to trancription cocktail and incubated at 40 °C for 2 hours, 4 °C for 1 minute.
|
| |
|
| Channel 2 |
| Source name |
Stratagene whole mouse reference RNA
|
| Organism |
Mus musculus |
| Characteristics |
sample type: reference
|
| Treatment protocol |
Tumors were harvested from bains of mice and immediately flash-frozen in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted s with Qiagen RNeasy kit according to manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
Labeling was performed according to the Low RNA Input Linear Amplification kit from Agilent. 1µg of RNA was primed with 1.2 µl of provided T7 promoter primer at 65 °C for 10 minutes. Reverse transcription was performed with provided Moloney Murine Leukemia Virus Reverse Transcriptase at 40 °C for 2 hours, 65 °C for 15 minutes, and 4 °C for 5 minutes. Laeled RNA was made by adding 1µl of 10mM Cy3 or Cy5 added to trancription cocktail and incubated at 40 °C for 2 hours, 4 °C for 1 minute.
|
| |
|
| |
| Hybridization protocol |
Labeled sample RNA and Stratagen Universal Mouse Reference RNA (Agilent, #740100) as a reference were cohybridized at 65 C for 17 hours in a rotating hybridization oven.
|
| Scan protocol |
Microarrays were scanned on an Agilent Technologies DNA Microarray Scanner with Surescan High-Resolution Technology (Part no. G2565CA)
|
| Data processing |
Image analyzed using Agilent Feature Extraction Software. Data was uploaded to the UNC microarray database (UMD). Data was normalized in UMD using Lowess normalization on the Cy3 and Cy5 channels. Data extraction was performed by selecting genes with an absolute signal intensity of at least 10 units in both dye channels and data present in at least 70% of experimental samples, afterwhich replicate probe IDs were collapsed by averaging. Samples from different batches were combined using the parametric settings in combatR, a module in the R statistical programming language (R Development Core Team, 2006 http://www.R-project.org) with no covariates included. Normalized data provided is the filtered, batch combined, non-median centered data used to draw conclusions in the study.
|
| |
|
| Submission date |
Jul 07, 2014 |
| Last update date |
Jan 31, 2016 |
| Contact name |
Ryan Miller |
| E-mail(s) |
miller@med.unc.edu
|
| Phone |
919-966-4333
|
| Organization name |
University of North Carolina
|
| Department |
Pathology
|
| Street address |
6109B NRB CB 7250
|
| City |
Chapel HIll |
| State/province |
North Carolina |
| ZIP/Postal code |
27599 |
| Country |
USA |
| |
|
| Platform ID |
GPL11202 |
| Series (1) |
| GSE59116 |
Core pathway mutations induce de-differentiation of murine astrocytes into glioblastoma stem cells that are sensitive to radiation but resistant to temozolomide. |
|
