normalization data transformation protocol
CEL files were generated from images (DAT files) using Command Console (Affymetrix, Santa Clara, CA), log2 RMA expression estimates were calculated using Partek Genomics Suite ID_REF = VALUE = log2 RMA expression estimate
array scanning protocol
GeneChips were scanned in a GeneAtlas Imaging Station (Affymetrix, Santa Clara, CA)
8 µg of cDNA was hybridized per microarray for 16 hours of hybridization at 45ºC, washing and staining of microarrays was performed using a GeneAtlas Fluidics Station (Affymetrix, Santa Clara, CA)
RNA was amplified for 23 cycles using the TransPlex Complete Whole Transcriptome Amplification Kit (Sigma; reference WTA2) and subsequently labeled using GeneChip Mapping 250K Nsp Assay Kit (Affymetrix; catalog # 900766), as described in Gonzalez Roca et al. 2012.
freshly isolated progenitor cells from lineage depleted bone marrow of ID2-GFP reporter were analysed. Cells were not cultivated prior RNA isolation.
nucleic acid extraction protocol
RNA was extracted using Agencourt RNAClean XP magnetic beads (Beckman Coulter) as described (Gonzalez Roca et al. 2010)
sample treatment protocol
cells were flow cytometrically isolated from lineage depleted bone marrow derived from ID2-GFP reporter mice and immediately lysed for RNA purification.