|
| Status |
Public on Jan 20, 2011 |
| Title |
epidermis1_4J/m2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
epidermis irradiated 4J/m2
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: epidermis
dose: 4J/m2
|
| Treatment protocol |
Irradiation with a solar simulated radiation five hours before recovering the samples
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Skin homogenisation was carried out with the Precellys 24 device (Bertin), using ceramic beads (CK14), in the presence of 350microlitres of RA1 buffer (Macherey-Nagel) supplemented with 3.5 microlitres of beta-mercaptoethanol. Th e device was set at a speed of 6300 rpm, with a cycle duration of 23 seconds and an interval time between 2 cycles of 2 minutes at 4°C. Six cycles were required for complete homogenisation. Tri-reagent (400 microlitres) was added followed by 150 microlitres of chloroform. The aqueous phase was recovered, mixed with 500 microlitres of 70% ethanol and transferred to a nucleospin RNA II colomn. RNA was recovered from the column following the manufacturer's instructions.
|
| Label |
Cy5
|
| Label protocol |
Total RNA (350ng) were amplified and labelled using a two color labelling protocol with the low input linear amplification kit according to the manufacturer's recommandations (Agilent)
|
| |
|
| Channel 2 |
| Source name |
Pool of all samples (control)
|
| Organism |
Homo sapiens |
| Characteristics |
reference: Pool of all samples
|
| Treatment protocol |
Irradiation with a solar simulated radiation five hours before recovering the samples
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Skin homogenisation was carried out with the Precellys 24 device (Bertin), using ceramic beads (CK14), in the presence of 350microlitres of RA1 buffer (Macherey-Nagel) supplemented with 3.5 microlitres of beta-mercaptoethanol. Th e device was set at a speed of 6300 rpm, with a cycle duration of 23 seconds and an interval time between 2 cycles of 2 minutes at 4°C. Six cycles were required for complete homogenisation. Tri-reagent (400 microlitres) was added followed by 150 microlitres of chloroform. The aqueous phase was recovered, mixed with 500 microlitres of 70% ethanol and transferred to a nucleospin RNA II colomn. RNA was recovered from the column following the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Total RNA (350ng) were amplified and labelled using a two color labelling protocol with the low input linear amplification kit according to the manufacturer's recommandations (Agilent)
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed using an Agilent oligonucleotide microarray in situ Hybridisation-Plus kit, following manufacturer's instructions.
|
| Scan protocol |
Scanned with th dynamic autofocus Agilent G2565A microarray scanner
|
| Description |
P20_4J_VS_Pool
|
| Data processing |
Agilent feature extraction software version 9.1 and the Bioconductor package Limma were used to extract and normalized the data.
Script under R (bioconductor with Limma package)
MA = normalizeWithinArrays(RGb,method=loess)
MA.q=normalizeBetweenArrays(MA,method=Gquantile)
[Data filtering criteria]
Flagged-signals excluded:
-Saturated in the 2 channels
-Non uniform
-SNR< 2,6 and not significantly different from local background
And only genes with less than 30% of missing values were selected
|
| |
|
| Submission date |
Jan 27, 2010 |
| Last update date |
Jan 20, 2011 |
| Contact name |
Nicolas Mouchet |
| E-mail(s) |
nicolas.mouchet@univ-rennes1.fr
|
| Organization name |
CNRS UMR6290
|
| Department |
Institut genetique et developpement de Rennes
|
| Lab |
Regulation transcriptomique et oncogenese
|
| Street address |
2 Av du Professeur Leon Bernard
|
| City |
Rennes |
| State/province |
Bretagne |
| ZIP/Postal code |
35043 |
| Country |
France |
| |
|
| Platform ID |
GPL1708 |
| Series (1) |
| GSE20062 |
In vivo identification of solar-responsive gene network |
|
