To evaluate the incidence of dentinal microcracks formation after root canal shaping procedures with HyFlex EDM and Vortex Blue rotary systems comparing with that of hand instrumentation using micro-computed tomography. Mandibular first molar teeth (n=60) having 50 to 200mesial root curvature were scanned using high resolution micro-CT imaging before root canal preparation to identify the presence of dentinal defects. Post-instrumentation cross-sectional images were taken and increased number and type of root defects were assessed and recorded. The data was statistically analysed by using chi-square test and McNemar?s test at a significance level of 5%. HyFlex EDM has shown greater increase in post instrumentation dentinal defects. Both rotary systems exhibited increased dentinal defects formation at coronal, middle regions which is statistically significant (P=0.042) when compared to apical region. HyFlex EDM has shown greater increase in post instrumentation dentinal defects followed by Vortex Blue rotary system and hand instrumentation resulted minimal defects.