|
| Status |
Public on Jan 31, 2018 |
| Title |
Hypothalamus ICI 5 mg/kg/day infusion Replicate 4 |
| Sample type |
RNA |
| |
|
| Source name |
Fetal Hypothalamus, ICI 182,780 5 mg/kg/day infusion
|
| Organism |
Ovis aries |
| Characteristics |
tissue: Fetal hypothalamus
developmental age: Around 130 days of fetal age
treatment: ICI 182,780 5 mg/kg/day
|
| Treatment protocol |
Chronically catheterized ovine fetuses were infused for 48 hours with: saline vehicle (Control, n=6), 17β-estradiol 500 ug/kg/day (Estradiol, n=4), ICI 182,780 5 μg/kg/day (ICI 5μg, n=4) and ICI 182,780 5mg/kg/day (ICI 5mg, n=5).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from fetal hyothalamus with Trizol (Life Technologies, Carlsbad, CA), and purified through Qiagen RNeasy+ kits with on-column DNase digestion (QIAGEN, Valencia, CA) according to manufacturers' protocols.Total RNA concentration was measured by Nanodrop ND-1000 and RNA quality was monitored by Agilent Bioanalyzer.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 500 ng RNA using the One-Color Quick Amp Labeling Kit (Agilent, Newcastle DE) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
600 ng of Cy3-labelled cRNA (specific activity: 10.22 to 12.38 pmol Cy3/µg RNA) was fragmented and hybridized to Agilent Sheep Oligo Microarrays (G4813A) design 019921 by the ICBR facility at the University of Florida according to Agilent's methodology.
|
| Scan protocol |
Slides were scanned on the Agilent DNA Microarray Scanner (G2505B) using one color (green) scan setting for 8x15k array slides, by the ICBR facility at University of Florida.
|
| Description |
Gene expression in fetal hypothalamus infused with 5 mg/kg/day of ICI 182,780-Replicate 4
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.1.1.1 (Agilent) using default parameters (protocol GE1-v5_10 and Grid: 019921_D_F_20100112) background detrended Processed Signal. The limma package was employed to import the raw data into R (http://www.r-project.org), perform background correction and normalize the data by the quantile normalization method. Control probes and low expressed probes were filtered out, retaining for further analysis the probes that were at least 10% brighter than the negative controls on at least four arrays.
|
| |
|
| Submission date |
May 31, 2017 |
| Last update date |
Jan 31, 2018 |
| Contact name |
Maria Belen Rabaglino |
| E-mail(s) |
m.b.rabaglino@uu.nl
|
| Organization name |
Utrecht University
|
| Department |
Population Health Science
|
| Street address |
Yalelaan 7
|
| City |
Utrecht |
| ZIP/Postal code |
3584 CL |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL14112 |
| Series (1) |
| GSE99497 |
Estradiol and ICI 182,780 infusions induce similar transcriptomic response in late gestation fetal hypothalamus |
|
