Sybr premix qrt pcr

Rats were euthanized by sevoflurane overdose at 24 h after SCII in accordance with the established protocol by the Experimental Animal Center of China Medical University. Segments L4-L6 of the spinal cord were collected to extract total RNA with the Trizol reagent (Takara, Otsu, Japan). The RNA was reverse-transcribed into cDNA using a Prime-Script RT reagent Kit with gDNA Eraser (Takara, Otsu, Japan) (Jia et al., 2019 (link)). The levels of miRNA were measured using an SYBR Premix qRT-PCR (Takara, Otsu, Japan) on the Applied Biosystems 7500 Real Time PCR system (Takara) with U6 as an internal control. The primer sequences are shown in Table 1. The 2^−ΔΔCt method was used to calculate the data.

Segments L4-L6 of the spinal cord at 48 h after SCII were collected to extract total RNA with using Trizol reagent (Takara, otsu, Japan). Then we used Prime-Script RT reagent Kit with gDNA Eraser (Takara) to reverse-transcribe RNA into cDNA using Prime-Script RT reagent Kit with gDNA Eraser (Takara). The mRNA, lncRNA and TF expression levels were determined using the SYBR PremixEx Taq II kit (Takara) with GAPDH as an internal control on Applied Biosystems 7500 real Time PCR system. The miRNA expression levels were determined using SYBR Premix qRT-PCR (Takara) on Applied Biosystems 7500 Real Time PCR system with U6 as an internal control (Santos et al., 2020 (link)). Table 1 demonstrated the primer sequences used in the present study. We calculated data through the 2^–ΔΔCt method.