|
| Status |
Public on Feb 05, 2014 |
| Title |
doxycycline replicate B, card B |
| Sample type |
RNA |
| |
|
| Source name |
MCF-10A Dox-on cells_test
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Doxycycline-inducible MCF-10A cells
treated with: 2μg/ml doxycycline for 48hr
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was harvested using the MicroRNeasy Mini Kit (Qiagen).
|
| Label |
FAM
|
| Label protocol |
Generation of cDNA and TaqMan qRT-PCR reactions were performed by the Dana-Farber Cancer Institute Molecular Diagnostics Laboratory as specified by the manufacturer. Briefly, 60ng of input RNA was reverse-transcribed using miRNA-specific stem-loop primers. Next, 12 cycles of pre-amplification reactions were performed using miRNA-specific forward primers and a universal reverse primer. Finally, the pre-amplification reactions were diluted, transferred to 384-well plates via microfluidic channels, and subjected to TaqMan miRNA amplification reactions using miRNA-specific FAM-labeled TaqMan probes.
|
| |
|
| Hybridization protocol |
n/a
|
| Scan protocol |
n/a
|
| Description |
SAMPLE 5
|
| Data processing |
Non-normalized data consist of raw Ct values that were reported for each miRNA assayed. Ct values above 30 are considered unreliable, according to the experience of the Molecular Diagnostics Laboratory, due to high intra-sample variability in this regime. Samples that did not show any amplification up to 40 cycles were labeled “Undetermined.” Normalized data were generated by subtracting the average Ct values of the housekeeping genes RNU44 and RNU48 in the sample from the Ct value of each miRNA. Fold-change data were calculated using the delta-delta Ct value method and are shown relative to untreated sample replicate A.
Normalized data report miRNA Ct values normalized against housekeeping genes (average Ct values of RNU44 and RNU48 for a particular sample). Fold-change data are relative to sample 1 (untreated condition, replicate A).
|
| |
|
| Submission date |
Feb 05, 2013 |
| Last update date |
Feb 05, 2014 |
| Contact name |
Michael Xiang |
| Organization name |
Dana-Farber Cancer Institute
|
| Department |
Medical Oncology
|
| Lab |
David A. Frank
|
| Street address |
44 Binney St
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL11316 |
| Series (1) |
| GSE44089 |
MicroRNA expression changes associated with specific STAT3 activation |
|
