Total RNAs of regions of interest were prepared with an RNeasy miniprep kit according to manufacturer's (Qiagen, Valencia, CA) protocol. RNA was quantified by using the NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE), and quality was assessed with a 2100 Bioanalyzer (Agilent Technologies, Palo Alta, CA) and with agarose gel electrophoresis. Depending on the brain region, between 1 and 5 µg of total RNA was used to prepare double-stranded cDNA with a cDNA synthesis kit (GIBCO/BRL) for first and second strand. Biotinylated cRNA was generated from these cDNAs with a BioArray high-yield RNA transcript labeling kit (Enzo Life Sciences). It was then purified with RNeasy spin columns (Qiagen) and fragmented before hybridization
Label
biotin
Label protocol
Approximately 1 to 5 µg of total RNA was processed to produce biotinylated cRNA targets.
Hybridization protocol
standard Affymetrix procedures
Scan protocol
standard Affymetrix procedures
Description
cocaine naive, septum, 0 hrs/day access to cocaine
