|
| Status |
Public on Jan 15, 2024 |
| Title |
MRC5_CSC_R3 |
| Sample type |
RNA |
| |
|
| Source name |
MRC5 fibroblast treated with CSC during 6 days
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Normal human fibroblast cells
|
| Treatment protocol |
MRC5 cells were cultured with DMSO or Cigaret Smoke Condensate (CSC) at 40µg/ml during 6 days
|
| Growth protocol |
MRC5 normal human fibroblasts (ATCC) were cultured in Dulbecco′s modified Eagle′s medium (DMEM, Life Technologies) with GlutaMax and supplemented with 10% FBS (Sigma-Aldrich) and 1% penicillin/streptomycin (ThermoFisher Scientific). Cells were maintained at 37°C under a 5% CO2 atmosphere. All experiments were carried out on cells at early passages (between 22 and 28).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from MRC5 cells using NucleoSpin® RNA (Macherey Nalgen) following the manufacturer's recommendations
|
| Label |
Cy3
|
| Label protocol |
cRNAs were synthesized and labeled with the Cy3 dye from 100 ng of total RNA using the one-color Low Input Quick Amp Labeling Kit (Agilent Technologies) following the manufacturer's recommandations, followed by RNeasy Mini column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. After fragmentation 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Mouse GE 4x44K v2 Microarrays (Agilent-026655) 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 briefly in acetonitrile.
|
| Scan protocol |
Whole Human Genome Microarrays 4x44K v2 (Agilent Technologies) were scanned with an Agilent DNA microarray scanner G2565CA (Agilent Technologies).Fluorescent signals were extracted and normalized with the Feature Extraction software version 10.5.1.1 (Agilent Technologies),
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities.
|
| |
|
| Submission date |
Jan 09, 2024 |
| Last update date |
Jan 15, 2024 |
| Contact name |
jean-michel flaman |
| E-mail(s) |
jean-michel.flaman@lyon.unicancer.fr
|
| Phone |
+33687477812
|
| Organization name |
Inserm 1052
|
| Department |
CRCL
|
| Street address |
28 rue Laennec
|
| City |
Lyon |
| ZIP/Postal code |
69373 |
| Country |
France |
| |
|
| Platform ID |
GPL13497 |
| Series (1) |
| GSE252801 |
Gene expression profiling of human fibroblast MRC5 cells exposed to cigarette smoke condensate (CSC) |
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