Anterior pituitary gland was harvested at the time of sacrifice, weighed and flash-frozen in liquid nitrogen. Tissue was homogenized in lysis buffer by passage through an 18- and then 22-guage needle. Total RNA was isolated using the Absolutely RNA Kit from Stratagene.
Label
biotin
Label protocol
Approximately 2 µg of total RNA was processed to produce biotinylated cRNA targets.
Hybridization protocol
standard Affymetrix procedures
Scan protocol
standard Affymetrix procedures
Description
ACI rats were obtained from Harlan (Indianapolis, IN). Empty silastic tubing implants were surgically inserted into control animals’ interscapular regions, as sham treatment, at 9 weeks of age. Each animal received an intraperitoneal injection of BrdU (50mg/kg) 4 hours prior to sacrifice. Animals were sacrificed at 21 weeks of age by decapitation and pituitary glands were immediately harvested, weighed and flash-frozen in liquid nitrogen. Tissues were homogenized through a 22-gauge needle and total RNA was extracted using the Absolutely RNA Kit (Stratagene). A total of 3 control males were used, resulting in two independent biological replicates.
Data processing
Affymetrix GeneChip Operating Software version 1.2
