GSE108503
Pathway analysis of RNA sequencing of various stages of myodifferentiation identifies conditions favoring type I and type II fibers, and highlights increased efficiency of combined differentiation
RNA-Seq of C2C12 cells differentiated for 0, 2, or 7 days using classic myogenic (DM1), adipogenic (DM2), or mixed (DM3) stimulation. Overall design: Fourteen samples were sequenced: two control samples at day 0 before differentiation was initiated, and 2 at each differentiation condition (DM1/DM2/DM3) at day 2 and day 7. cDNA libraries were constructed using stranded protocol from New England Biolabs. Poly-A selection was used to remove the majority of rRNA from samples. Each library was sequenced with single-end 50 bp reads on HiSeq 2500 sequencer, generating between 25 and 40 million raw reads. Reads were aligned to primary assembly of mouse genome GRCm38 using Gencode vM12 annotation, using STAR v.2.5.1b. Overall alignment rate for each sample was in 95-99% range, with 74-83% of original reads aligning uniquely, and less than 5.5% rRNA contamination. Aligned reads were then quantified using RSEM (v.1.2.31) and analyzed using DESeq2 and fGSEA R packages, as well as MsigDB and Morpheus online tools.