Abstract: BACKGROUND: Our objective in this study was to assess the association between eNOS gene, that achieves synthesis of nitric oxide especially in the endothelial cells known to have an important role in angiogenesis and vasculogenesis, G894T, intron 4 VNTR (27-bp repeat) and T786C functional polymorphisms and retinopathy of prematurity (ROP), which is an important cause of morbidity in premature or low birth weight babies. METHODS: A total of 139 babies who were followed up in our neonatal intensive care unit because of premature birth in our hospital or admitted to our unit. 69 of them had retinopathy of prematurity and comprised the patients group. The remaining 70 babies who did not have ROP comprised the control group. An additional of 1 ml of blood samples were drawn from babies who were in the study groups during routine laboratory analysis. eNOS gene polymorphisms were determined by using polymerase chain reaction method. RESULTS: eNOS G894T, intron 4 VNTR and T786C gene polymorphisms did not differ between the patient and control groups (p > 0.05). Using logistic regression analysis; while gender did not differ between two groups; gestational age, birth weight, time on mechanical ventilation differ between two groups. After adjustment for variables other than eNOS gene polymorphisms, we found no significant difference in the genotype distribution of eNOS G894T, intron 4 VNTR and T786C polymorphisms (p > 0.05). CONCLUSION: We observed no association between ROP and eNOS gene polymorphisms but needs more investigation.