|
| Status |
Public on Feb 01, 2014 |
| Title |
09 13344 |
| Sample type |
RNA |
| |
|
| Source name |
tumor tissue
|
| Organism |
Homo sapiens |
| Characteristics |
disease state: breast cancer tumor
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The frozen normal and tumor tissues were collected for RNA extraction. Total RNA was isolated with the MirVana kit (Ambion, Austin, TX)
|
| Label |
Cy3
|
| Label protocol |
500ng of total RNA from each sample was dephosphorylated with Calf Intestinal Alkaline Phosphatase (CIP) at 37ºC for 30’ following a denaturalization and then a ligation for 2h at 16ºC using Agilent miRNA Complete Labeling and Hyb Kit (Agilent Technologies; Palo Alto, CA). In this step a molecule of Cyanine3-pCp is incorporated to the 3’ end of RNA molecules.
|
| |
|
| Hybridization protocol |
Labeled RNA was dried and resuspended with Hybridization Buffer and Blocking Agent, incubated 10’ at 100ºC and transfered to an ice water bath for 5’. Samples were hybridized in a volume of 45ul to the Human miRNA V2 Oligo Microarray (Agilent) for 20 hours at 55°C and 20rpm. Microarrays were then washed at room temperature for 5min in Gene Expression Wash Buffer 1 and 5min at 37ºC in Gene Expression Wash Buffer 2 (Agilent Technologies; Palo Alto, CA).
|
| Scan protocol |
Arrays were scanned on an Agilent G2565BA microarray scanner under default settings recommended by Agilent Technologies for miRNA microarrays with 100% PMT and 5μm resolution.
|
| Data processing |
Data was extracted using Agilent Feature Extraction Software (Agilent Technologies; Palo Alto, CA). Extracted Log2-transformed intensities were quantile normalized to make all data comparable.
|
| |
|
| Submission date |
Feb 06, 2013 |
| Last update date |
Feb 01, 2014 |
| Contact name |
Andrea Feliciano |
| E-mail(s) |
andrea.feliciano@vhir.org
|
| Organization name |
Vall d´Hebron Research Institute
|
| Lab |
Oncology and Molecular Pathology
|
| Street address |
Passeig Vall d´Hebron 119-129
|
| City |
Barcelona |
| ZIP/Postal code |
08035 |
| Country |
Spain |
| |
|
| Platform ID |
GPL14767 |
| Series (1) |
| GSE44124 |
profile miRNA expression in breast cancer |
|
