Species of the genus Ganoderma are important medicinal and edible mushrooms, with polysaccharides being its main active ingredient. The submerged fermentation of Ganoderma is an important source of polysaccharides in the mycelia. In this study, a single factor test and an orthogonal test were employed to optimize the medium and fermentation conditions with the objective of obtaining a high content of polysaccharide in the liquid submerged fermentation culture mycelia of Ganoderma species. The results demonstrated that the optimal culture medium formula for polysaccharide accumulation in mycelia was: soluble starch, 25 g/L; wheat bran, 3 g/L; and KH2PO4, 4.5 g/L, with a pH of 4.0, and the optimal culture conditions were a temperature of 27°C, a rotational speed of 90 r/min, and an incubation period of 11 d. Furthermore, the mycelia of 20 strains from nine species of Ganoderma were cultured under optimal fermentation conditions, and the content of polysaccharide in mycelia and its antioxidant activity were analyzed. The findings revealed that the polysaccharide content in the mycelia of strains GD and Du996 was notably elevated, at 56.97 and 53.22 mg/g, respectively. The polysaccharide content was found to be high in the mycelia of G. sichuanense and G. lucidum, with an average of 35 mg/g. Nevertheless, no notable discrepancy was observed in the polysaccharide content of the mycelia of the cultivated and wild strains. Moreover, the antioxidant activity analysis revealed that the DPPH clearance rate of strains Du1320, Du280, and GD was 92.85%, 90.86%, and 89.06%, respectively. The ABTS clearance rate of strain GSD was the highest (75.39%), and the hydroxyl radical clearance rate of strain GD was the highest (81.98%). In conclusion, this study optimized the medium and conditions for high polysaccharide yield in the liquid submerged fermentation culture of Ganoderma and identified the strains with the highest yield and antioxidant activity. The present results may provide a theoretical basis for the future production of polysaccharides in mycelia by liquid fermentation of Ganoderma as well as for the development and utilization of strains with high polysaccharide yield and antioxidant activity.