Clearly defining mineral deformation and slip systems is crucial for an in-depth analysis of the intrinsic mechanisms governing mineral responses to external stress and temperature, as well as their rheological weakening processes. The rapid advancement of science and technology and its deep integration into the geological field provide an opportunity for a detailed analysis of structural deformation behavior and mechanisms. In this study, quartz and amphibole from representative naturally deformed rocks were used as examples. Based on microstructural analysis, a comprehensive assessment was conducted using a substantial dataset of mineral lattice preferred orientation measurements obtained via an electron backscatter diffraction (EBSD) probe mounted on a field-emission scanning electron microscope. By examining microstructural features, EBSD mapping data, dislocation geometry types, and properties, a detailed analytical method for grain boundary trace and misorientation axes was developed. The results reveal that the strain adjustment and grain refinement process in quartz occur mainly through the {m}<a> slip system, dominated by the subgrain rotational recrystallization mechanism in quartz veins. It was also found that in mylonitic amphibolites, amphibole porphyroclasts exhibit pronounced fine-grained deformation behavior, primarily driven by subgrain rotational recrystallization. Furthermore, amphibole undergoes multi-slip system interactions, predominantly governed by the [001] direction through dislocation creep in banded amphibolites. Thus, integrating EBSD grain boundary trace analysis with misorientation axis analysis and microstructural characterization enables a comprehensive determination of microgeological information—including composition, shape, grain size, orientation, boundaries, and strain—of deformed minerals. This approach further elucidates the evolution of orientation from the grain interior to intergranular regions (or matrix). Moreover, the dominant slip system in mineral deformation processes can be effectively defined and correlated with the deformation environment, which has substantial geological implications.