Transcription profiling of Theiler stage 17 wild type mouse anterior vs posterior mouse sclerotome

This experiment was performed in order to identify transcriptional differences between the anterior- and posterior-halves of mouse sclerotome. Cells-derived from the anterior- and posterior-sclerotome-halves from maturing mouse somites were compared to identify transcripts that are differentially expressed between these two halves of the somite. c57b6-/- female mice were time-mated, and at 9.5-dpc embryos were harvested. Theiler stage 17 embryos were selected for dissection. A stripe of segmented paraxial mesoderm corresponding to somites SX-SXVIII (standard somite nomenclature) was dissected from one-side of an embryo. The most anterior-third of up to 7 sclerotomes within each sample were dissected and pooled in RNALater. RNA was isolated using RNeasy Micro RNA extraction spin columns (Qiagen). Reverse transcription and initial cDNA amplification was performed using 18 PCR cycles with the SMART™ cDNA system (Clontech). Amplified cDNA was then re-amplified and labeled using the GeneChip® IVT Labeling Kit (Affymetrix).