Quantitative PCR for Friedreich's Ataxia

Polymerase chain reaction (PCR) conditions to measure GAA•TTC repeat length were described previously.^{21 (link)} Quantitative reverse transcriptase PCR (qRT-PCR) was performed using the qScript One-Step SYBR Green qRT-PCR kit from Quanta Biosciences (Gaithersburg, MD) according to the manufacturer’s instructions. qRT-PCR reactions were detected on a PTC-200 thermal cycler with the Chromo4 real-time module (MJ Research, Waltham, MA). Primers to detect FXN mRNA were previously described.^{6 (link)} Primers for neuronal characterization were as follows: MAP2-R1 (5′-CAGGAGTGATGGCAGTAGAC-3′), MAP2-F2 (5′-TTTGGAGAGCATGGGTCAC-3′) for the MAP2 gene, HUC-F1 (5′-GGTTCGGGACAAGATCACAG-3′), and HUC-R1 (5′-CTGAACTGGGTCTGGCATAG-3′) for the ELAVL3 gene. qRT-PCR primers for pluripotency mRNA markers were as previously published.^{20 (link)} Neuronal cell gene expression profiling was performed on Illumina (San Diego, CA) HT12 arrays, and statistical analysis was as described in Ku et al.^{20 (link)}