Quantifying Gene Expression in ALS Mice

Total RNA was isolated from TA muscle of WT and hSOD1^G93A mice using RNeasy Lipid Tissue extraction kit (QIAGEN Inc., Alameda, CA, USA) per the manufacturer’s protocol. The total RNA was purified from genomic DNA contamination using Turbo DNAse treatment (Ambion, Life Technologies, Carlsbad, CA, USA) then converted to cDNA by means of a reverse transcription kit (Agilent Technologies Inc., Santa Clara, CA, USA) per the manufacturer’s protocol. Commercially available gene-specific Taqman probes (Applied Biosystems, Life Technologies, Carlsbad, CA, USA) were used to amplify target gene of interest as described previously [8 (link)]. Relative target gene expression to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was determined using this formula: 2^-∆CT where ∆Ct = (Ct target gene – Ct GAPDH) [20 (link)]. Final measures are presented as relative levels of gene expression in hSOD1^G93A mice compared with expression in WT controls.