Abstract: To investigate the effects of substituting dietary soybean meal with Chlorella on growth performance, muscle quality, and intestinal microbiota of mirror carp (Cyprinus carpio var. specularis), five experimental diets were formulated by replacing dietary 0, 25%, 50%, 75%, and 100% of soybean meal with Chlorella, which named CV0 (Control), CV25, CV50, CV75, and CV100, respectively. The mirror carp with an initial body weight of (20.00±0.01) g were fed five experimental diets for 60d. The results showed that compared with the CV0, the final body weight in CV50 and CV75 groups, the weight gain rate in CV50, CV75 and CV100 groups, the specific growth rate and lipid retention efficiency in CV25, CV50, CV75 and CV100 groups, as well as the protein retention efficiency in CV25, CV50 and CV75 groups were significantly increased (P<0.05). The activities of α-amylase, trypsin and lipase in the intestine of CV50 group were significantly higher than those in the CV0 (P<0.05). Meanwhile, the villus height and muscularis thickness of intestine CV25 and CV50 groups were also significantly increased (P<0.05). In terms of health status, replacing dietary 50% soybean meal with Chlorella significantly increased the T-AOC levels and SOD activity in both liver and intestine (P<0.05), while also markedly reducing liver MDA level (P<0.05). Meanwhile, intestinal CAT activity was seriously enhanced in the CV25 and CV50 groups (P<0.05). Compared with the control group, the CV50 group showed obviously increased serum alkaline phosphatase levels, while serum alanine aminotransferase and aspartate aminotransferase levels were significantly declined (P<0.05). Pathological damage was observed in both liver and kidney tissues in the CV100 group. In terms of muscle quality, substituting 50% dietary soybean meal with Chlorella significantly improved the nutritional value of fish by increasing the crude protein and crude lipid level in both whole body and muscle, as well as the muscle NEAA, EAA, and PUFA level (P<0.05). Principal component analysis of muscle texture characteristics showed that the CV50 group exhibited superior texture properties compared to other groups, indicating that replacing dietary 50% soybean meal with Chlorella improved muscle texture. Additionally, replacing 50%, 75%, and 100% of soybean meal with Chlorella increased muscle redness and yellowness (P<0.05). The 50% dietary soybean meal was substituted by Chlorella elevated the relative abundance of dominant intestinal microbiota (Proteobacteria and Verrucomicrobiota). The quadratic regression analysis based on growth performance indicated that the optimal replacement level of soybean meal with Chlorella was 58.14%—70.54%. In conclusion, complete replacement of soybean meal with Chlorella had no negative effects on the growth performance of mirror carp, but caused damage to liver and kidney tissues. However, appropriate replacement of soybean meal with Chlorella promoted the growth, improved muscle quality of mirror carp, with the optimal replacement level being 58.14%—70.54%.