Pharyngeal Satellite Cell Expression Profile

To compare the differential gene expression between sorted satellite cells collected from pharyngeal verses limb muscle. Myogenic cells from pharyngeal and limb (gastrocnemius and quadriceps) muscles were isolated, sorted using FACS, and pooled into samples numbering 150,000-200,000 cells collected from 10-30 mice per experiment. Samples were sent to the Emory University Integrated Genomics Core facility for total RNA isolation using Qiagen miRNEAsy kit with on-column DNAse treatment followed by one round of amplification using NuGEN’s WT-Ovation Pico amplification kit. Analysis of genomic gene expression was performed using an Illumina Mouse WG-6 v2.0 Expression BeadChip. Data was extracted using the Illumina HiScan Scanner and iScan control software. Illumina Genome Studio 2011.1 software suite was used to normalize probe level intensity data with background correction using manifest MouseWG-6_V2_0_R1_11278593_A.txt. Detection p-values were calculated as the proportion of negative control probes with expression greater than the regular probe in question using Partek Genome Studio. Data were further analyzed using both MetaCore Genego (https://portal.genego.com; Thomson Reuters) and Gene Set Enrichment Analysis (www.broadinstitute.org/gsea/index.jsp; Broad Institute).