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Abstract The purpose of the present study was to compare the effect of reciprocation motion (WaveOne system) and continuous rotation motion (ProTaper system) versus hand instrumentation (K-FlexoFile) on root canal transportation and centering ability using Endodontic cube which is a modification of Bramante et al technique. Thirty mesiobuccal canals of extracted human permanent lower molars with angles of curvature ranging from 25 to 40 degrees were selected for this study. Samples were divided into three equal groups according to the type of the instrument used for preparation. The roots were mounted in the endodontic cube with clear self-curing casting resin then cross-sectioned at 4 mm, 6 mm and 8 mm levels from the apex by using a diamond 0.3mm thick cutting disc. Pre-instrumented cross-sections were photographed under a stereomicroscope then reassembled again by an external muffle for instrumentation. The final apical preparation diameter was size 25 for all experimental groups. The cross-sections were photographed again. The images of the pre- and postinstrumented canals were superimposed using Adobe Photoshop. Canal transportation and centering ratio were measured and calculated by using a modification of Gambill et al method together with Image J program. Results revealed that the overall performance of WaveOne showed better centering ratio and statistically significant less canal transportation than both K-FlexoFiles and ProTaper system. There was no statistically significant difference between the mean values of the tested groups at the different levels regarding the canal transportation and centering ratio. The apical level of the three groups showed lower mean values of canal transportation followed by the middle level then the coronal level, with statistically significant difference between the three levels of K-FlexoFile group |