Objective: The aim of this study was to compare the effect of four instruments used as orifice openers on the cervical dentine thickness of human mandibular first molars, using cone beam computed tomography (CBCT). Materials and methods: Forty human permanent mandibular first molars were decoronated and sectioned using a water-cooled safe sided diamond disc. The mesial roots were vertically mounted in a transparent acrylic resin box before CBCT scanning. Sections were scanned at 2.0, 3.0 and 4.0 mm below the cervical line. The samples were randomly divided into four equal groups according to the orifice opener used (n = 10). The canals were instrumented using Sendoline, Pre-RaCe, Hyflex and ProTaper files. The root canals were scanned after the cervical flaring using the same pre-flaring scanning protocol; cervical dentine thickness was measured before and after flaring. One way analysis of variance ANOVA was used to compare between the tested groups and each thickness. Tukey's post hoc test was used for pairwise comparison between the means. A dependent t-test was used for intra-group comparison (P ≤ 0.05). Results: For the Hyflex group, the percentage change in distal cervical thickness was significantly higher than the mesial one at the level of 2.0 mm below the cervical line. At the level of 3.0 mm and 4.0 mm for the Sendoline group, the percentage change in distal cervical thickness was significantly higher than the mesial one. Conclusion: None of the rotary instruments used in this study caused any damage to the root canal walls. Keywords: cervical flaring, cone-beam computed tomography (CBCT), dentine thickness, orifice opener, root canal instruments