Band electrode submerged arc overlay welding was used to weld welding strip of EQ309L stainless steel on the surface of Q345R matrix. The microstructure of EQ309L side, Q345R side, and their interfaces were observed by electron backscattered diffraction (EBSD) technique. Results show that most grains in Q345R matrix are seriously distorted, and the banded grains along the original rolling direction can be clearly observed. The average grain size of Q345R matrix is 30~40 μm in the overheated coarse-grain region, and the grain coarsening is not severe. The average grain size of the fine-grain region is 10~20 μm in the Q345R matrix. The transition zone is 35~40 μm away from the fusion line of Q345R matrix, and it has body-centered cubic (bcc) structure. The EQ309L layer has coarse grains, showing the microstructure of columnar crystal with obvious <100> texture.