|
| Status |
Public on Apr 01, 2017 |
| Title |
PC3ER_10nM beta-estradiol (E2)_24hrs |
| Sample type |
RNA |
| |
|
| Source name |
PC3ER, 10nM beta-estradiol (E2), 24hrs
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: PC3ER (derived from prostate cancer cell line PC3)
agent: E2
time: 24 hrs
|
| Treatment protocol |
10 nM of beta-estradiol (E2) was added to the medium.
|
| Growth protocol |
Cells were maintained in DMEM containing 10% of activated-charcoal treated FBS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
QIAGEN RNeasy mini kit was used.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 ug of Cy3-labelled cRNA (specific activity >9.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55ul containing 25x Agilent fragmentation buffer and 10x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 ul of 2x GEx Hybridization buffer HI-RPM (Agilent) was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays 4x44k (G4112F) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then rinsed with Acetonitrile for cleaning up and drying.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 4x44k array slides.
|
| Description |
PC3ER E2 treatment
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 9.1.3.1 (Agilent) using default parameters (protocol GE1-v5_91_0806 and Grid: 014850_D_20060725) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
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| |
|
| Submission date |
Apr 20, 2011 |
| Last update date |
Apr 01, 2017 |
| Contact name |
Shingo Inaguma |
| Organization name |
Aichi Medical University School of Medicine
|
| Department |
Department of Pathology and Promoting Center for Clinical Research
|
| Street address |
Nagakute
|
| City |
Aichi |
| ZIP/Postal code |
480-1195 |
| Country |
Japan |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE28900 |
Identification of GLI1 direct and indirect target genes in human prostate cancer cell line PC3 |
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