|
| Status |
Public on Jul 23, 2013 |
| Title |
UESCs_48h_replicate1 |
| Sample type |
RNA |
| |
|
| Source name |
UESCs, 48h, synchronization
|
| Organism |
Rattus norvegicus |
| Characteristics |
cell type: uterus endometrial stromal cells
|
| Treatment protocol |
The uterus endometrial stromal cells were isolated from Per2-dLuc transgenic rats on day 4.50 of gestation. The harvested cells were seeded onto 35 mm collagen-coated dishes at the density of 2×105 cells/dish with 2 mL of culture medium (phenol red-free DMEM/F12 supplemented with 10% charcoal-treated FBS and 1× PS). Then, cells were cultured in serum-free medium supplemented with 1× antibiotic–antimycotic, 1× Insulin-Transferrin-Selenium, 0.1% bovine serum albumin, and 100 nM progesterone for additional 2 days prior to other treatments. The cultured UESCs were synchronized with 100 nM dexamethasone for 2 h in the serum-free medium. RNA samples isolated from cultured cells at 30, 36, 42, and 48 h after dexamethasone synchronization.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using the Versagene Blood RNA Purification kit (Gentra Systems, Minneapolis MN) following the manufacturer's recommendations. The protocol includes differential lysis of red and white blood cells, and an on-column DNase digestion. Globin message was further reduced using GLOBINclear (Ambion Inc., Austin, TX) to specifically remove both a- and b- globin. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
cRNA was amplified and labelled using a Low input Quick Amp Labelling Kit (Agilent Technologies).
|
| |
|
| Hybridization protocol |
cRNA was hybridized to a 44K 60-mer oligomicroarray (Whole Rat Genome Microarray Kit v3 ; Agilent Technologies) according to the manufacturer's instructions.
|
| Scan protocol |
The hybridized microarray slides were scanned using an Agilent scanner. The relative hybridization intensities and background hybridization values were calculated using Feature Extraction Software version 9.5.1.1 (Agilent Technologies).
|
| Description |
Gene expression after dexamethasone synchronization
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 9.5.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. The raw signal intensities and flags for each probe were calculated from the hybridization intensities and spot information according to the procedures recommended by Agilent Technologies using the Flag criteria in the GeneSpring Software. In addition, the raw signal intensities of two samples were log2-transformed and normalized by the quantile algorithm with the Bioconductor.
|
| |
|
| Submission date |
Jul 22, 2013 |
| Last update date |
Jul 23, 2013 |
| Contact name |
MASA-AKI HATTORI |
| E-mail(s) |
[email protected]
|
| Organization name |
Kyushu University
|
| Department |
Graduate School of Agriculture
|
| Lab |
Laboratory of Reproductive Physiology
|
| Street address |
6-10-1, Hakozaki, Higashi-ku
|
| City |
Fukuoka |
| ZIP/Postal code |
812-8581 |
| Country |
Japan |
| |
|
| Platform ID |
GPL14746 |
| Series (1) |
| GSE49080 |
Profiling of circadian genes expressed in the uterus endometrial stromal cells of pregnant rats as revealed by DNA microarray coupled with RNA interference |
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