Total colonic RNA was extracted by using TRIzol (Vazyme Biotech Co., Ltd) and subsequently purified through lithium chloride precipitation [19 (link)]. Real-time quantitative PCR (RT-PCR) was performed with 2 × ChamQ SYBR Color qPCR Master Mix (Vazyme Biotech Co., Ltd.) following the manufacturer’s protocols. β-actin was used as the housekeeping gene for colonic tissue. The quantification of mRNA levels was measured using the threshold cycle (2−ΔΔCT) method as described previously [20 (link)]. The primers are shown in Supplementary Table 1 .
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