|
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 |
|