Expression data from three layers in the growth plate

The developing vertebrate growth plates in long bones have three morphologically distinct layers of chondrocytes: Round Cell layer, Flat Cell layer, and Hypertrophic Cell layer. Round chondrocytes differentiate into flat chondrocytes, and then flat chondrocytes differentiate into hypertrophic chondrocytes. To investigate the genetic programs underlying the chondrocyte differentiation, we obtained total RNA from the microdissected cryosections from each layer and then performed microarray analysis. We euthanized 3 litters at birth to obtain biological triplicate data. Using cryosections of unfixed fresh frozen hind limbs, we manually dissected each layer on the glass slide under a dissecting microscope. We collected each piece of the sections in lysis buffer and stored at -80°C prior to RNA extraction. To prepare one sample, we dissected 18 to 20 tibiae from the same litter.