Aim: To evaluate and compare the remaining dentin thickness post-instrumentation of teeth with three rotary systems using CBCT. Materials and methods: In this study, a total of 36 extracted human mandibular premolars were taken. All teeth were decoronated till the cementoenamel junction. Then they were randomly divided into 3 groups according to the system used in root canal preparations namely Group I: TruNatomy file system, Group II: ProTaperNext file system, and Group III: Hyflex EDM file system. Conventional endodontic access was prepared in each tooth, and the working length was determined using a 15-k file. Pre-operative cone beam computed tomography (CBCT) scan of the teeth was taken. Cleaning and shaping of the canal were done according to the file system. Post instrumentation CBCT scan of teeth was taken. The pre-and post-instrumentation scanned images of the teeth were assessed, the difference in the mesiodistal and buccolingual area was calculated and the statistical difference was analyzed. Results: The TruNatomy files showed the least dentin removal in all regions compared to the other two files. Conclusion: All three groups under study showed statistically significant differences in dentin thickness post-instrumentation. However, The TruNatomy File System showed better conservation of tooth structure compared to ProTaperNext And Hyflex EDM file system.