|
| Status |
Public on Nov 02, 2020 |
| Title |
Ileum_001 |
| Sample type |
RNA |
| |
|
| Source name |
Ileum, Zn treatment 15 days_replicate 1
|
| Organism |
Mus musculus |
| Characteristics |
tissue: ileum
gender: female
age: 7 weeks
diet: 15 days of Zn supplemented diet
|
| Treatment protocol |
Three groups of mice (n=4) were randomized to receive distinct doses of ZnCl2 solutions (Sigma–Aldrich, St. Quentin–Fallavier, France) administered in the drinking water (Zn: 250 mg per kg body weight (mg/kg), 50 or 12 mg/kg) while a control group consisted in standard uncontaminated drinking water (n=4). Zn concentration is negligible in drinking water and its concentration is 25 mg/kg in standard chow (Diet A04C-10, Scientific Animal Food and Engineering, Augy, France). After 15 days of Zn treatment, mice are sacrificed and sections (0.5 /1cm in length) from the median colon and terminal ileum were collected and processed with RNA stabilization solution (RNA-later, Ambion, Life Technologies).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Mouse colon and ileum samples were homogenized using the FastPrep instrument (MP Biomedicals), total RNA was isolated using RNAspin columns (Macherey-Nagel).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.2 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS 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.
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| |
|
| Hybridization protocol |
1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug 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 Agilent Mus musculus Sure Print GE 4x44 v2 microarrays with oligonucleotide 45,220 probes (G2519F) 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 by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Scanner GenePix 4200B (Molecular Device) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5um).
|
| Data processing |
The scanned images were analyzed with GenePix Pro Software 6.0 (Molecular Device) using default parameters 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 |
Nov 04, 2019 |
| Last update date |
Nov 03, 2020 |
| Contact name |
Anca Lucau-Danila |
| Organization name |
University of Lille
|
| Department |
Biology
|
| Lab |
Charles Viollette Institute
|
| Street address |
Cité Scientifique SN2 303
|
| City |
Villeneuve d'Ascq |
| ZIP/Postal code |
59655 |
| Country |
France |
| |
|
| Platform ID |
GPL11202 |
| Series (1) |
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