|
| Status |
Public on Jul 29, 2016 |
| Title |
Hamster_Control_Sample |
| Sample type |
RNA |
| |
|
| Source name |
ATCC CCL-49 (RPMI 1826)
|
| Organism |
Mesocricetus auratus |
| Characteristics |
hybrid: : Mesocricetus auratus
original tumor type: Melanoma
sample type: Cultured monolayer
|
| Treatment protocol |
Tissue samples were preserved as formalin-fixed paraffin-embedded (FFPE) tissue blocks for up to 40 years and sectioned to 5-10 um thick for nucleic acid isolation.
|
| Growth protocol |
Various human tumors were inidividually grafted to the cheek pouches of adult, unconditioned golden hamsters (Mesocricetus auratus), and the resulting tumor for each was serially passaged in hamsters for multiple years.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy FFPE Kit (Qiagen, Germantown, MD) according to manufacturer’s instructions (3h incubation at 55 degrees C followed by glass fiber filtration). RNA purity was checked by OD260/280 readings and RNA integrity was checked using Agilent Bioanalyzer capillary electrophoresis profiles.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cDNA from each sample weas prepared by conversion of 50 ng total RNA to cDNA by reverse transcription, amplification by the SPIA method, and fragmentation and labeling with biotin using the Ovation Pico WTA System v2 and Encore Biotin Module kits according to the manufacturer’s instructions (NuGEN, San Carlos CA).
|
| |
|
| Hybridization protocol |
5.5 ug of fragmented, biotin-labeled cDNA were hybridized for 16 hr at 45 degrees C on GeneChip Human U133_X3P arrays (Affymetrix; catalog no. 900516). GeneChips were washed and stained with streptavidin-phycoerythrin using the Affymetrix Fluidics Station 450, according to the manufacturer's recommended protocol.
|
| Scan protocol |
GeneChips were scanned using the Affymetrix GCS 3000 7G.
|
| Description |
ATCC CCL-49 (RPMI 1826)
|
| Data processing |
Fluorescent signals were collected and processed by AGCC and MAS 5.0 using Affymetrix default analysis settings and global scaling as the normalization method in Expression Colsole. The trimmed mean target intensity (TGT) of each array was set to 250. Following MAS 5.0 normalization and global scaling, unannotated probe sets and probe sets for which no signal value was greater than the median signal for AFFX spike-in controls with all Absent Detection Calls were omitted from further analysis. Subsequently, because an intact hamster cell line (sample IMM006, cell line CCL-49) control RNA sample was used for comparison with the four human-hamster hybrid FFPE samples, all remaining signal values for the hamster cell line sample were multiplied by the ratio of the median signal in all FFPE hybrid samples for AFFX spike-in control probe sets called present in all samples divided by the median signal for the same probe sets in the hamster control sample.
|
| |
|
| Submission date |
Jun 06, 2014 |
| Last update date |
Jul 29, 2016 |
| Contact name |
David M Goldenberg |
| E-mail(s) |
dmg.gscancer@att.net
|
| Organization name |
Garden State Cancer Center
|
| Department |
Center for Molecular Medicine and Immunology
|
| Street address |
300 The American Road
|
| City |
Morris Plains |
| State/province |
NJ |
| ZIP/Postal code |
07950 |
| Country |
USA |
| |
|
| Platform ID |
GPL1352 |
| Series (1) |
| GSE58277 |
In-Vivo Fusion of Human Cancer and Hamster Stromal Cells Permanently Transduces and Transcribes Human DNA |
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