|
| Status |
Public on Feb 01, 2016 |
| Title |
control (culture media) |
| Sample type |
RNA |
| |
|
| Source name |
Skin biopsy
|
| Organism |
Tursiops truncatus |
| Characteristics |
location: Tuscany shore, Italy
gender: female
tissue: Skin
cultured in: control media
|
| Treatment protocol |
Distinct slices were separately incubated for 24 h at room temperature (ranging from 24 to 28 °C) in cell culture media containing BPA (0.1 µg/ml or 1 µg/ml), or PFOA (0.1 µg/ml or 1 µg/ml), or the vehicle (ethanol for BPA and methanol for PFOA).
|
| Growth protocol |
Slices (about 2 mm-thick) spanning the epidermis and dermis were cut from skin samples of the stranded specimen immediately after collection, to set up the organotypic cultures
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the Aurum TM Total Fatty and Fibrous Tissue kit (Bio-Rad) following the manufacturer’s instructions.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Amplification kit (Agilent) 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 and the Agilent 2100 Bioanalyzer.
|
| |
|
| Hybridization protocol |
500ng of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to custom Tursiops truncatus Microarray (Agilent) 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 dried immediately with a wash in acetonitrile (Agilent).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner using one color scan setting for 4x44k array slides (Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
TT_SKIN_CNT
Gene expression wild dolphin skin
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software A8.5.3 (Agilent) using default parameters (protocol GE1) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Jun 16, 2015 |
| Last update date |
Feb 01, 2016 |
| Contact name |
Annalaura Mancia |
| E-mail(s) |
annalaura.mancia@unife.it
|
| Organization name |
University of Ferrara
|
| Department |
Life Sciences and Biotechnology
|
| Street address |
via L.Borsari, 46
|
| City |
Ferrara |
| ZIP/Postal code |
44121 |
| Country |
Italy |
| |
|
| Platform ID |
GPL17696 |
| Series (1) |
| GSE69917 |
TRANSCRIPTOMIC ANALYSIS OF BOTTLENOSE DOLPHIN (Tursiops truncatus) SKIN BIOPSIES TO ASSESS THE EFFECTS OF EMERGING CONTAMINANTS ON A TOP PREDATOR IN THE MEDITERRANEAN SEA |
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