Background: men’s participation is crucial for promoting of breastfeeding. There are a few studies on the effectiveness of father’s education on the nursery performance. Therefore, this study aims to assess the mothers’ performance following father’s education on breastfeeding. Method: This randomized clinical trial was performed on 76 fathers in two groups of control (38 men) and intervention (38 men) in a selected health center in Iran. They were spouses of primiparous women; fathers of a healthy, single neonates. Two groups of intervention and control were selected using randomized sampling method. The tools for data collection were: a “Demographic and Maternal-Infant Information Questionnaire”, and a “Fathers’ support for Breastfeeding Assessment Tool”, a “Checklist to Assess Mothers’ Breastfeeding Performance” and an “Exclusive Breastfeeding Questionnaire”. The intervention was: fathers’ education about “breastfeeding” and “the ways of support including encouragement and planning for breastfeeding”. The education was performed in the intervention group in two sessions with one-week interval. Fathers in the control group did not receive the education. Mothers’ breastfeeding performance, father’s support for breastfeeding and exclusive breastfeeding was compared before and after intervention, and between groups after four months’ intervention. Data was analyzed using SPSS 26. Results: Between groups comparison showed, before intervention, the mean scores for breastfeeding performance was not significantly different between two groups (P=0.7). However, after the intervention, it was higher in the interventional group comparing to the control group (T test; p<0.001). Intra-groups comparison demonstrated, after intervention, the improvement was not seen after 4 months in the control group (p=0.6), while there was a significant improvement in the intervention group comparing to before intervention (Paired test; p<0.001). Conclusion: The results showed that father’s education improves breastfeeding performance of nursery mothers. Since, breastfeeding is an important factor for maternal-infant health, providing fathers’ participation education to promote breastfeeding programs leads to long and effective exclusive breastfeeding.Trial Registration: IRCT201508248801N10. Registered 31 August 2016.