This work is meant to study the effect of galacto-oligosaccharide (GOS) intervention in the fecal microbiota of obese mice on lipid metabolism and anti-obesity in mice. A pseudo-germ-free mice model was established by administration of a mixture of antibiotics. The fecal microbiota of GOS-fed obese mice are transplanted into pseudo-germ-free mice. To investigate the effects of GOS intervention on lipid metabolism, lipid quadruple, bile acid metabolism, and intestinal microbiota in obese mice. At the same time, the expression of BAS synthase and the effects of FXR-SHP and FXR-FGF15-FGFR4 signaling pathways on BAS homeostasis were also explored. The fecal microbiota of obese mice intervened by GOS could reshape the structure of intestinal microbiota, regulate the synthesis of bile acids in the enterohepatic circulation and the expression of transport-related factors, and promote metabolic ability to exert anti-obesity effects.