To investigate the relationship between the number of viable rhizobia Mesorhizobium huakuii cells inoculated and the nodule formation and nitrogen fixation in Astragalus sinicus, two inoculation methods—seed surface and root inoculations—were employed, using liquid bacterial agents as inoculants. Through a sterile sand-pot experiment system, the minimum number of rhizobia required for nodule formation and nitrogen fixation in A. sinicus was determined. The results showed that when seed surface inoculation was applied, a rhizobial concentration of 102 cfu/seed led to the greening of plant leaves. The fresh weight and height of the above-ground parts of the plants significantly increased compared to treatments with lower inoculation doses. Meanwhile, nitrogen-fixing nodules developed on the roots of the A. sinicus plants, demonstrating a positive growth-promoting effect. When plant root inoculation was conducted, a single A. sinicus plant required only an inoculation dose of 101 cfu/plant to initiate nodule formation and nitrogen fixation. Notably, regardless of the inoculation method used, when the inoculation dose reached 103 cfu/plant per A. sinicus plant, the nodule formation and nitrogen fixation as well as plant growth, were significantly better than those in the low-inoculation groups. Further increasing the inoculation dose resulted in a stabilization of the symbiotic nitrogen fixation phenotype. To fully ensure the nodule formation and nitrogen fixation of rhizobial inoculants after application in the field, and taking into account the economic effectiveness of inoculant application and the complexity of soil conditions, this study has determined that 103 cfu/plant is the minimum viable cells for inoculating A. sinicus with M. huakuii rhizobia.