This study investigated the effect of two lipid sources on reproduction performance and growth in pearl gourami. For this purpose, 180 fish (3.32 ± 0.25 g) were fed with three isoenergetic (19.80) and isonitrogenous diets (480 g/kg protein) including FO (80 g/kg fish oil), FS (40 g/kg fish oil and 40 g/kg soybean oil) and SO (80 g/kg soybean oil) for 10 weeks before maturation. At the end of the trial, there was no significant difference in weight gain, feed conversation ratio and body composition between fish fed FO and FS diets. Individuals fed dietary FO had significantly higher levels of n‐3 long‐chain polyunsaturated fatty acids in the muscle (130.5 g/kg lipid) and ovary (140.4 g/kg lipid) as compared with those fed SO diet (64.5, 103.6 g/kg, respectively) (p  < .05). Feeding pearl gourami with FO and FS diets enhanced regarding absolute fecundity, relative fecundity, the fertilization rate, larvae total length and survival at 3 day posthatch (p  < .05). Also, 17 beta‐estradiol in plasma of fish fed dietary FO (6.2 ng/L) was higher than those fed SO diet (1.7 ng/L) (p  < .05). In conclusion, we suggest FS diet for broodstock nutrition of pearl gourami as a model for asynchronous multi‐batch spawning fish.