The aim of this ex vivo study was to evaluate the vertical-fracture resistance of roots obturated with a newly developed tricalcium silicate cement (BioRoot RCS; Septodont, Saint Maur Des Fosses, France) using cold lateral compaction technique (LC) or matched-taper single-cone gutta-percha technique (SC). Decoronated 82 single-rooted mandibular premolars were chemo-mechanically prepared and then randomly divided into 6 experimental groups (n = 12) and 2 control groups (n = 5): Group 1 iRoot SP-LC; Group 2 iRoot SP-SC; Group 3 MTA Fillapex-LC; Group 4 MTA Fillapex-SC; Group 5 BioRoot RCS-LC and Group 6 BioRoot RCS-SC. In the positive-control group, roots were instrumented but not filled, and in the negative-control group, roots were neither instrumented nor filled. All samples were incubated for two weeks and then subjected to vertical loading force (1 mm/min) until fracture. The force required to fracture each specimen were determined, and the data were statistically analyzed. The highest significant fracture resistance was recorded for the iRoot SP-LC, iRoot SP-SC, BioRoot RCS-LC, and BioRoot RCS-SC groups, with no significant difference among them (p > 0.05) when compared with the positive-control group (p < 0.05), whereas the lowest significant values were observed in the MTA Fillapex-LC and MTA Fillapex-SC groups (p < 0.05). The vertical fracture resistance of roots obturated with BioRoot RCS and iRoot SP sealers using either LC or SC technique was found to be similar to that of intact teeth. BioRoot RCS, newly developed tricalcium silicate cement, might have the potential to reinforce the instrumented teeth against vertical root fracture.