Several studies have indicated that feeding female animal with fatty acids could result in alteration of offspring sex ratio; however, there is little information available regarding the effect of feeding male animal with fatty acids on sex ratio of offspring. The aim of current study was to investigate the effect of paternal feeding with n-3 and n-6 polyunsaturated fatty acids (PUFAs) on sperm characteristics and sex ratio of offspring in mice. Male mice received 2.50% of palm oil (control), fish oil (n-3 PUFAs) or sunflower oil (n-6 PUFAs) for two months. Sperm kinematics and viability were assessed using computer-assisted sperm analyzer and eosin-Y staining, respectively. Additionally, female mice were randomly introduced to males of three experimental groups to determine reproductive parameters including litter size, conception rate and sex ratio. Fish oil enhanced sperm concentration, motility and viability, whereas sunflower oil decreased sperm concentration. Nevertheless, progressive motility, velocity, linearity and straightness of sperms were not affected by source of fatty acids. Although supplementation with fish oil resulted in male-biased sex ratio, palm and sunflower oils did not impact sex ratio. Besides, source of fatty acids failed to influence conception rate and litter size. In conclusion, the present study provided evidence for the impact of paternal fish oil consumption on sex ratio of offspring for the first time. Moreover, it showed beneficial effect of n-3 PUFAs and adverse effects of n-6 PUFAs on seminal parameters.