Tract-based spatial statistics (TBSS)6 is a software pipeline widely employed in comparative analysis of the white matter integrity from diffusion tensor imaging (DTI) datasets. In this study, we seek to evaluate the relationship between different methods of atlas registration for use with TBSS and different measurements of DTI (fractional anisotropy, FA, axial diffusivity, AD, radial diffusivity, RD, and medial diffusivity, MD). To do so, we have developed a novel tool that builds on existing diffusion atlas building software, integrating it into an adapted version of TBSS called DAB-TBSS (DTI Atlas Builder-Tract-Based Spatial Statistics) by using the advanced registration offered in DTI Atlas Builder7. To compare the effectiveness of these two versions of TBSS, we also propose a framework for simulating population differences for diffusion tensor imaging data, providing a more substantive means of empirically comparing DTI group analysis programs such as TBSS. In this study, we used 33 diffusion tensor imaging datasets and simulated group-wise changes in this data by increasing, in three different simulations, the principal eigenvalue (directly altering AD), the second and third eigenvalues (RD), and all three eigenvalues (MD) in the genu, the right uncinate fasciculus, and the left IFO. Additionally, we assessed the benefits of comparing the tensors directly using a functional analysis of diffusion tensor tract statistics (FADTTS10). Our results indicate comparable levels of FA-based detection between DAB-TBSS and TBSS, with standard TBSS registration reporting a higher rate of false positives in other measurements of DTI. Within the simulated changes investigated here, this study suggests that the use of DTI Atlas Builder’s registration enhances TBSS group-based studies.