Abstract <p>Pain following endodontic treatment is a common complication potentially linked with genetic polymorphisms that modulate pain biomarkers. Hence, this study aimed to explore the association between pain experienced after endodontic treatment and genetic polymorphisms in the genes encoding inducible nitric oxide synthase (<i>NOS2</i>), and suppressor of cytokine signaling (<i>SOCS1</i>). The study involved 108 participants with single-rooted teeth, single canals, and necrotic pulp related to asymptomatic apical periodontitis. All endodontic treatments were executed in a single session. Pain and tenderness levels were measured using a visual analog scale (VAS) on the 1st, 2nd, 3rd, 4th, 5th, 6th, 7th, 14th, and 30th day post-treatment. Genomic deoxyribonucleic acid (DNA) was extracted from saliva cells and the genetic polymorphisms rs2779249, rs2297518, rs243327 and rs33977706 were genotyped using real-time polymerase chain reaction. Genotype distribution was assessed using univariate and multivariate Poisson regressions through generalized estimating equations (GEE) and categorized based on the presence or absence of pain and tenderness, and the significance threshold was set at <i>p</i> &lt; 0.05. The genetic polymorphism rs2297518 in the <i>NOS2</i> gene was associated with pain in the recessive model (<i>p</i> = 0.019) and to tenderness in both the codominant (<i>p</i> = 0.008) and recessive (<i>p</i> &lt; 0.001) models. The genetic polymorphisms rs2779249 in <i>NOS2</i> and rs243327 and rs33977706 in <i>SOCS1</i> showed no association with pain or tenderness (<i>p</i> &gt; 0.05). In conclusion, the genetic polymorphism rs2297518 in the <i>NOS2</i> gene was associated with pain after root canal treatment. These findings contribute to a better understanding of the genetic factors influencing postoperative pain in endodontics, which may help in developing personalized pain management strategies and improving patient care.</p> Graphical Abstract <p></p>