|
| Status |
Public on Jun 02, 2009 |
| Title |
Epidermis_Raf_30 Days 4OHT_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
epidermis, 30 days Raf activation, +4OHT
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Regenerated human epidermis
recipient strain: CB-17 scid/scid
|
| Treatment protocol |
Tissue was excised and frozen immediately in OCT on dry ice and stored at -20C from weeks to months. Prior to LCM, tissue was sectioned (6um) and stored at -20C for up to 4 days. Sections were stained with cresyl-violet (LCM staining kit, Ambion) and immediately processed by laser capture microdissection. Slides were not allowed to sit more than 2.5 hours at room temp for processing.
|
| Growth protocol |
Epidermal tissue co-expressing ER:H-RasG12V and IkBaM was regenerated on female scid/scid mouse recipients. Grafts were allowed to heal for at least 3 weeks before Ras activation via daily i.p. injections of 730ug of 4OHT (in 110ul of a corn oil/ethanol mixture). Duplicate grafts were harvested after 0, 5, 20, and 35 days of 4OHT treatment.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from laser capture microdissected tissue using RNAqueous kit following manufacturer's instruction (Ambion)
|
| Label |
biotin
|
| Label protocol |
Total RNA was amplified and fragmented using Message Amp II-Biotin following manufacturer's instruction (Ambion). 100-200ng of total RNA was amplified using 16 hr IVT reaction.
|
| |
|
| Hybridization protocol |
Samples were hybridized by the Stanford P.A.N. facility using a Fluidics Station 450 and GeneChip Hybridization Oven 640 following manufacturer's instruction (GeneChip Expression Analysis Technical Manual Rev.2, Affymetrix). For the fluidics protocol, a Midi_euk2v3_450 script was used.
|
| Scan protocol |
Samples were scanned by the Stanford P.A.N. facility a on Affymetrix Scanner 3000 G7 (default target signal set at 500). Also, for the fluidics protocol, we used Midi_euk2v3_450.
|
| Description |
Regenerated human epidermis
Raf(+4OHT)_A
|
| Data processing |
The raw cel files were imported into GeneSpring 7 (Agilent) and RMA normalized. The only other normalization that was applied was setting values less than 0.01 to 0.01.
|
| |
|
| Submission date |
Mar 19, 2009 |
| Last update date |
May 14, 2009 |
| Contact name |
Paul A Khavari |
| Organization name |
Stanford University
|
| Department |
Dermatology
|
| Lab |
Khavari Lab
|
| Street address |
259 Campus Dr
|
| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305-5168 |
| Country |
USA |
| |
|
| Platform ID |
GPL571 |
| Series (1) |
| GSE15299 |
Modeling Inducible Human Tissue Neoplasia Identifies an ECM Interaction Network Involved in Cancer Progression |
|
