RNA was isolated from whole brain and liver by a commercial RNA extraction kit (RNAXEL, Eurobio), treated with Dnase and cleaned using RNeasy columns (Qiagen). Equal amount of total RNA was pooled from 3 mice within each of the 6 experimental groups (3 strains, 2 conditions = 6 ; in triplicate : 6 * 3 = 18 chips) for both brain and liver ( 36 chips in total).
Label
biotin
Label protocol
Target preparation were performed as described by the Affymetrix GeneChip Expression Analysis Technical Manual (http://www.affymetrix.com), and were conducted at the Genomics Platform of the NCCR Frontiers in Genetics at the University of Geneva (Switzerland) . Equal quantities of total RNA from three individual mice of each strain were pooled. We generated a hybridization mixture containing 15 ug of biotinylated cRNA according to the protocols provided by Affymetrix.
Hybridization protocol
Fifteen micrograms of biotinylated cRNA from each sample were fragmented and hybridized to GeneChip Mouse 430 2.0 arrays, according to standard procedures. Washing and staining was performed using the EuGE-WS2v5 protocol.
Scan protocol
Scanning was done on an Affymetrix GeneChip Scanner 7G.
Description
none
Data processing
Normalized expression signals were calculated from Affymetrix CEL files using RMA (Irizarry et al., 2003).
