OBJECTIVES: To investigate the renoprotective effects of Gegen Qinlian Decoction (GQD) in Diabetes mellitus (DM) rats via the intestinal barrier and microbiota. METHODS: GQD was analyzed by UPLC. STZ-induced DM rat models and antibiotic-induced sterile DM rat models were established, and fecal microbiota transplantation was performed in the latter. Renal function, oxidative stress, serum inflammatory factors, and pathological alterations were assessed. Intestinal cells and tight junction were observed by transmission electron microscopy. Inflammatory factors in the colon and tight junction protein expression were evaluated. The gut microbiota and its abundance were assessed by 16sRNA sequencing. KEY FINDINGS: Four components were determined in the GQD, including puerarin, baicalin, berberine, and liquiritin. After GQD treatment, Scr and BUN were reduced, renal pathological changes were attenuated, intestinal cell swelling was reduced, intestinal tight junctions were improved, and GQD modulated the intestinal microbiota. Furthermore, a fecal bacterial solution containing GQD reduced renal lesions, improved intestinal tight junctions, and regulated intestinal microbiota in DM rats. CONCLUSIONS: GQD regulated the intestinal microbiota of DM rats, reduced intestinal inflammation, and repaired the intestinal barrier, thus reducing the burden on the kidneys, and exerting a protective effect on the kidneys of DM rats.