Ribospin 2 kit

Total RNA was extracted from cells that were pretreated with SA (100, 200, and 400 μM) followed by treatment with 50 μM of 6-OHDA using the Ribospin™ II kit (GeneAll Biotechnology Co., LTD, Seoul, Korea). After measuring the total RNA concentration using a UV/Vis Nano Spectrophotometer (MicroDigital Co., Ltd., Sungnam, Korea), 1 μg of total RNA from each group was reverse transcribed to synthesize cDNA using the GoScript Reverse Transcription System Kit (Promega Co., Madison, WI, USA). qRT-PCR was performed using a TaKara Thermal Cycler Dice Real-Time System (Takara Bio. Inc., Shiga, Japan) using the TOPreal™ qPCR 2× PreMIX Kit (Enzynomics Co., Daejeon, Korea). The primer sequences are as shown in Table 1.

Total RNA was isolated from the cells treated with 500 μM H₂O₂ alone or pretreated with 0.1, 0.5, and 1 μM Cu I followed by treatment of 500 μM H₂O₂ for 24 h by using a Ribospin™ II kit (GeneAll Biotechnology Co., LTD, Seoul, Korea). To examine the expression levels of mitochondrial biogenesis genes, 1 μg total RNA from the cells in each group was reverse transcribed into cDNA using ImProm II reverse transcriptase (Promega Co., Madison, WI, USA) with oligo-dT priming. qRT-PCR was performed using a Takara Thermal Cycler Dice Real-Time System (Takara Bio. Inc., Shiga, Japan) with SYBR Green (Takara) as a fluorescent dye. The primer sequences were as follows: peroxisome proliferator-activated receptor α (PPARα), forward 5′-GGC AAT GCA CTG AAC ATC GAG-3′ and reverse 5′-GCC GAA TAG TTC GCC GAA AG-3′; peroxisome proliferator-activated receptor γ coactivator- (PGC-) 1β, forward 5′-GTG AGA TAG TCG AGT GCC AGG TG-3′ and reverse 5′-TTC TCA GGG TAG CGC CGT TC-3′; estrogen-related receptor α (ERRα), forward 5′-GCT GAA AGC TCT GGC CCT TG-3′ and reverse 5′-TGC TCC ACA GCC TCA GCA T-3′; nuclear respiratory factor- (NRF-) 1, forward 5′-CAC TCT GGC TGA AGC CAC CTT AC-3′ and reverse 5′-TCA CGG CTT TGC TGA TGG TC-3′; and 18S, forward 5′-TTC TGG CCA ACG GTC TAG ACA AC-3′ and reverse 5′-CCA GTG GTC TTG GTG TGC TGA-3′.