Sn-Ag solders are widely used for advanced electronic packaging. The combined effect of Ag element and temperature gradient (TG) on the formation of Sn grains in Cu/Sn-xAg/Cu micro solder joints was elucidated systematically. Numerous small-sized β-Sn grains were formed in both Cu/Sn/Cu and Cu/Sn-0.5Ag/Cu micro solder joints after reflow with or without TG. The Cu/Sn-0.5Ag/Cu joint was found to have more twinning β-Sn structures. The formation of the multiple β-Sn grains in these two joints was attributed to the presence of multiple tetrahedral metastable short-range order (SRO) structures which acted as nuclei for the nucleation and growth of β-Sn. The existence of TG slightly enhanced the preferred orientation characteristics of Sn grains. For the joints with Ag content was or higher than 2 wt.%, several Sn grains were formed without TG, while a single or highly oriented Sn grains were observed with TG. The number and orientation of Sn grains were affected by the combined effect of Ag element and TG significantly. The β-Sn grains formed without TG were based on the {101} type cyclic twinning configuration clusters that stabilized by Ag atoms and acted as nuclei. The formation of a single or highly preferred β-Sn grains was benefit from the combined effect of Ag element and TG. The results provide theoretical guidance for optimizing the composition of Sn-Ag solders and controlling the microstructure of the joints, thereby contributing to the advancement of electronic packaging technologies.