SH-SY5Y Cell RNA Extraction and qPCR

SH-SY5Y cells were used for total RNA extraction, which was performed by using the RNAqueous ^®-4PCR kit (Ambion Inc., Austin, TX, USA) as previously reported [99 (link)]. Before cDNA synthesis, the integrity of RNA was evaluated by denaturing electrophoresis in TAE 1.2% agarose gel. cDNA was synthesized using 1 μg of total RNA for all samples using a High-Capacity cDNA Reverse Transcription kit (Applied Biosystems, Foster City, CA, USA) under the following conditions: 50 °C for 2 min, 95 °C for 10 min, 95 °C for 15 s, and 60 °C for 1 min for 40 cycles. RTq-PCR was performed using Master Mix TaqMan^®Gene Expression and the 7.500 RT-PCR instrument (Applied Biosystems), targeting genes in the Taqman Array 96-Well Plate (P/N: 4414292): brain-derived neurotrophic factor (BDNF, Hs02718934_s1), glial cell-derived neurotrophic factor (GDNF, Hs01931883_s1), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, Hs99999905_m1).