The purpose of this study was to investigate the dissimilar joint properties of S45C carbon steel with multi-layer and multi-pass welding. We used four weld rods labelled A, B, C, and D to experimentally examine the influence on microstructures. A, B, and C were medium carbon steels while D was stainless steel. The microstructures of joint beads were observed with an optical microscope. The specimens for tensile tests were made in accordance with American Society for Testing and Materials (ASTM) E8M. The experimental results were as follows: (1) Pearlite structures and bainite structures were formed in the welded zone by the weld rods of medium carbon steels. The yield strength and ultimate strength of the specimens were 360 and 460 MPa, respectively. (2) The feature of welded specimen D was close to stainless steel because the stainless steel rod included some Cr and Ni. The yield strength and ultimate strength of specimen D showed increases of 35 and 65% compared to the other specimens. In addition, a ferritic-austenitic stainless steel, known as dual phase stainless steel, was observed in the optical microscope.