2.8 qPCR
Total RNA of colonic tissues and cells were extracted by using Trizol (Accurate Biotechnology, Hubei, China) accroding to manufacturer’s instructions. The cDNA was synthesized by using High Capacity cDNA Reverse Transcription Kit (ThermoFisher Scientific, NY, USA) and amplified with SYBR Green Master Mix (Thermo Fisher Scientific, NY, Massachusetts, USA). The folding changes of mRNA expression of target gene were normalized to GAPDH by ΔΔCt method. The unit of the variable is determined as the folded mean of the control.