|
| Status |
Public on Dec 01, 2011 |
| Title |
pre-miR-27b 2 |
| Sample type |
RNA |
| |
|
| Source name |
Prostate cancer LNCaP cells
|
| Organism |
Homo sapiens |
| Characteristics |
transfection: pre-miR-27b
experiment: 1
batch: 2
cell line: LNCaP
|
| Treatment protocol |
Pre-miRNA and control oligonucleotides (Ambion, Austin, TX) at 30-nM final concentration were transfected using Lipofectamine 2000 (Invitrogen, Carlsbad, CA), as directed by the manufacturer. Briefly, cells were plated in six-well plates at 2.5X10^5 cells per well, grown for 48-72 hr, and then switched to antibiotic free media prior to transfection. Next, 1.5 µl oligos at 50µM and 4 µl Lipofectamine 2000 were added individually to 250 µl of Opti-MEM (Invitrogen) and then combined and incubated for at least 20 minutes, before being added to cells (2.5 ml media in total). Cells were collected at 24 hr for microarray studies.
|
| Growth protocol |
Cells were plated in six-well plates at 2.5x105 cells per well, allowed to grow for 48-72 hours and then switched to antibiotic free media before being transfected.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TRIZOL reagent (Invitrogen, Carlsbad, CA) as directed by the manufacturer. The quantity and quality of RNA was assessed with a Nanodrop 1000 spectrophotometer (Thermo Scientific, Waltham, MA) and Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA).
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol (Expression Analysis Technical Manual, Affymetrix).
|
| |
|
| Hybridization protocol |
GeneChip® Human Genome U133A 2.0 (Affymetrix Inc., Santa Clara, CA) arrays were used following standard Affymetrix procedures as outlined in the Affymetrix technical manual. GeneChips® were washed and stained using the Affymetrix Fluidics Station 450.
|
| Scan protocol |
GeneChips® were scanned using the Affymetrix GeneChip ® Scanner Scanner 3000
|
| Description |
Gene expression data from LNCaP cells transfected with pre-miR oligos
|
| Data processing |
Microarray data (CEL files) were normalized using RMA and MAS.5 filtered for genes with their detection calls as “P” or “M” within at least 3 samples.
|
| |
|
| Submission date |
Aug 23, 2011 |
| Last update date |
Dec 01, 2011 |
| Contact name |
Robert Scott Hudson |
| Organization name |
NIH
|
| Department |
NCI
|
| Lab |
The Laboratory of Human Carcinogenesis
|
| Street address |
National Cancer Institute Room Blgd.37/3044
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20852 |
| Country |
USA |
| |
|
| Platform ID |
GPL571 |
| Series (1) |
| GSE31620 |
MicroRNA-1 is a candidate tumor suppressor and prognostic marker |
|
