|
| Status |
Public on Sep 26, 2018 |
| Title |
Non-lesional_Skin substitute_M49 |
| Sample type |
RNA |
| |
|
| Source name |
Psoriatic skin substitute
|
| Organism |
Homo sapiens |
| Characteristics |
condition: Non-lesional
Sex: Male
age: 49
|
| Treatment protocol |
Biopsies of skin substitute were taken after 21 days of culture at the air-liquid interface.
|
| Growth protocol |
Fibroblasts were cultured in Dulbecco-Vogt modification of Eagle's medium (DMEM) supplemented with 10% bovine growth serum, 100UI/mL penicillin and 25μg/mL gentamicin. Keratinocytes were cultured in a combination of DMEM with Ham's F12 (3:1), supplemented with 5% Fetal Clone II serum, 5μg/mL insulin, 0.4μg/mL hydrocortisone, 1e-10M cholera toxin, 10ng/mL human epidermal growth factors, 100UI/mL penicillin and 25 μg/mL gentamicin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the RNeasy Mini Kit (QIAGEN, Cat#74104).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 50-200 ng RNA using the One-Color LowInput QuickAmp Labeling Kit One-Color (Agilent) according to the manufacturer's instructions
|
| |
|
| Hybridization protocol |
600ng of Cy3-labelled cRNA (specific activity >8.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 25 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole SurePrint G3 Human GE 8x60K (G4851A) 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.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent SureScanner (G4900DA) using one color scan setting for 1x60k array slides (Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10,7 (Agilent) 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.
|
| |
|
| Submission date |
Sep 25, 2018 |
| Last update date |
Sep 26, 2018 |
| Contact name |
Roxane Pouliot |
| E-mail(s) |
roxane.pouliot@pha.ulaval.ca
|
| Phone |
418-656-2131 7042
|
| Organization name |
Centre de recherche en organogenese expérimentale de l'Université Laval/LOEX
|
| Street address |
1401, 18ème rue, aile R
|
| City |
Québec |
| State/province |
Québec |
| ZIP/Postal code |
G1J 1Z4 |
| Country |
Canada |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE120464 |
The Tissue-Engineered Human Psoriatic Skin Substitute: A Valuable In Vitro Model to Identify Genes with Altered Expression in Lesional Psoriasis |
|
