1 ug of total RNA was used to transcribe DIG-labeled cRNA using Applied Biosystems Chemiluminescent RT-IVT Kit V2.0.
Hybridization protocol
Microarray hybridization (using twenty micrograms of fragmented, DIG-labeled cRNA) and processing were performed according to Applied Biosystems protocols.
Scan protocol
Chemiluminescence detection, imaging, auto gridding, and image analysis was done according to Applied Biosystems protocols and the 1700 Chemiluminescent Microarray Analyzer Software v. 1.0.3.
Description
The mouse cortex samples were obtained from P1 C57/B6 mice (n = 19), which were purchased from Charles River Laboratories (Charles River Laboratories, Inc., Willmington, MA).The mouse cortex was used as a reference sample. Universal mRNA reference was still being developed when the study started, so it was not utilized. All samples were placed in TRIzol reagent (Invitrogen, Carlsbad, CA) and stored at -80ºC before RNA extraction. Total RNA was extracted with TRIzol reagent according to the manufacturer's instructions. For a quality control measure of the samples, total RNA was ran through a 1% agarose gel and using the 2100 Agilent BioAnalyzer System (Agilent Technologies, Palo Alto, CA) to check for possible contamination and degradation.
Data processing
Signal intensities across microarrays were normalized using the quantile normalization (www.bioconductor.org).
