Background: One of the causative factors of root defects is the increased friction produced by rotary instrumentation. A high canal curvature may increase stress, making the tooth more susceptible to dentinal cracks. The purpose of this study was to evaluate dentinal micro-crack formation with the ProTaper NEXT and ProTaper Universal systems using LED transillumination, and to analyze the micro-crack generated at the point of maximum canal curvature. Material and Methods: 60 human mandibular premolars with curvatures between 30?49° and radii between 2?4 mm were used. The root canals were instrumented using the Protaper Universal® and Protaper NEXT® systems, with the aid of the Proglider® system. The obtained samples were sectioned transversely before subsequent analysis with LED transillumination at 2 mm and 8 mm from the apex and at the point of maximum canal curvature. Defects were scored: 0 for no defects; and 1 for micro-cracks. Results: Root defects were not observed in the control group. The ProTaper NEXT system caused fewer defects (16.7%) than the ProTaper Universal system (40%) ( P <0.05). The ProTaper Universal system caused significantly more micro-cracks at the point of maximum canal curvature than the ProTaper NEXT system ( P <0.05). Conclusions: Rotary instrumentation systems often generate root defects, but the ProTaper NEXT system generated fewer dentinal defects than the ProTaper Universal system. A higher prevalence of defects was found at the point of maximum curvature in the ProTaper Universal group.