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Differentiated instruction (DI) is a teaching approach that aims to achieve learning for diverse students. This study explores the impact of DI-focused strategies in a course in Teacher Education on STEM teacher candidates’ (TCs’) DI understandings and implementation. The research questions are: 1) How do TCs develop the curriculum to include DI using digital video games (DVGs)? 2) To what extent are DVGs effective tools to implement DI? The analysis of eight DVGs, developed by the TCs, shows the course effectiveness in promoting TCs’ conceptions and implementation of DI. Furthermore, DVGs are effective tools to differentiate instruction by facilitating pacing variation for different students, integrating multimodalities, utilizing engaging features, representing different learners, promoting conceptual understanding, and enabling different assessment forms.