Making gradient in the microstructure of metals is proven to be a superior method for improving their mechanical properties. In this research, we 3D print, characterize and evaluate the mechanical properties of 316L Stainless Steel with a gradient in their microstructure. During 3D printing, the gradient in the microstructure is created by tailoring the processing parameters (hatch spacing, scanning speed, and laser power and scanning speed) of the Selective Laser Melting (SLM). The Materials with Graded Microstructure (MGMs) are characterized by optical and scanning electron microscopy (SEM). Image processing framework is utilized to reveal the distribution of cells and melt pools shapes and sizes in the volume of the material when the processing parameters change. It is shown that the laser power, scanning speed and the hatch spacing have a more significant effect on the size and shape of cells and melt pools compared to the speed. Multiple Dog bones are 3D printed with a microstructure that has smaller features (cells and melt polls) at the edges of the structure compared to the center. Tensile and fatigue tests are performed and compared for samples with constant and graded microstructures.