Quantitative Real-Time PCR

RNA was extracted from tissues using RNA Isolation kit (Thermo Fisher Scientific). After evolution of quantity and quality, its concentration was normalized to 15 ng/µL. Reverse transcription was performed with High-Capacity Reverse Transcription Kit (Life Technologies, Grand Island, NY, USA). A qPCR 96-well reaction plate was used for the process with TaqMan (Life Technologies) primers and probes for BDNF (Rn02531967_s1), according to the manufacturer’s protocol, on Applied Biosystems^® 7500 Fast Real-Time PCR Instrument (Applied Biosystems, Foster City, CA, USA). Gapdh (Rn01775763_g1) and Tbp (Rn01455646_m1) were selected as endogenous control genes based on the pilot experiment. Relative expression was calculated using the ΔΔCq method.