Mg₂(Si,Sn)-based thermoelectric materials, which are environmentally friendly and low-cost, have great development potential in a moderate temperature range. Electronic transport properties of Mg₂Si_1-xSn_x alloys can be optimized by doping elements. Doping is still one of the most effective methods of optimizing electronic transport performance, such as carrier concentration, mobility, and effective mass. The most effective doping elements are Sb and Bi. Much attention has been paid to the influence of element type and doping content. Different substitution sites will also greatly affect the electronic transport parameters. In this work, the defect formation energy value of Mg₂Si_0.375Sn_0.625 alloy for substituting Sb atoms and Bi atoms for Sn sties and Si sites, respectively, are calculated by first-principles calculations. The influence on electronic transport parameters is systematically analyzed by combining the calculated results of band structures and density of states. Corresponding component Sb and Bi atoms doped Mg₂Si_0.375Sn_0.625 alloys are prepared by rapid solidification method, and microstructures, Seebeck coefficients, and electrical conductivities of the alloys are measured. Combined with the predicted results by solving the Boltzmann transport equation, electronic transport performances are compared and analyzed. The results indicate that both Sn and Si sites are equally susceptible to Sb and Bi doping, but the Si sites are preferentially substituted due to their lower ∆E_f values. Doped Bi atoms provide a higher electron concentration, and Sb atoms offer higher carrier effective mass. Thus, the maximum σ value of the Mg₂Si_0.375Sn_0.615Bi_0.01 alloy is 1620 S/cm. The maximum S value of the Mg₂Si_0.365Sn_0.625Sb_0.01 alloy is –228 μV/K. Correspondingly, the highest PF value for this alloy is 4.49 mW/(m·K) at T = 800 K because of the dominant role of S values. Although its power factor is slightly lower, the Mg₂Si_0.375Sn_0.615Sb_0.01 alloy is expected to exhibit lower lattice thermal conductivity due to the lattice shrinkage caused by substituting Sb sites for Sn sites. The optimal doping concentration of the Bi-doped alloy is lower than that of the Sb-doped alloy. These results are expected to provide a significant reference for optimizing the experimental performance of Mg₂(Si, Sn)-based alloys.