Omega‐3 long‐chain polyunsaturated fatty acid (n‐3 LC‐PUFA) increases in aquatic products contributes to improving meat quality, thereby positively impacting human health. Different from marine fish which primarily obtain n‐3 LC‐PUFAs directly from zooplankton and algae, freshwater fish mainly utilize dietary linolenic acid (ALA) as a substrate to synthesize n‐3 LC‐PUFAs. Our team has successfully created a transgenic rapeseed oil (TRO) with high ALA content. Therefore, we here assessed the impacts of four different diets (LR, low‐fat rapeseed oil (RO) diet; HR, high‐fat RO diet; LTR, low‐fat TRO diet; HTR, high‐fat TRO diet) on growth performance, lipid accumulation, fatty acid composition, antioxidant capacity, immunity and serum biochemical indexes of juvenile largemouth bass (Micropterus salmoides), an economically valuable freshwater fish. The results showed no significant difference in survival rate among the four dietary groups. No significant differences in body weight gain and final weight were found between the LR and LTR groups, as well as between HR and HTR groups. No matter if it was a high‐fat or low‐fat diet, compared with the RO diet, TRO diets significantly increased the content of n‐3 LC‐PUFA, improved meat quality, effectively alleviated lipid accumulation in livers and muscles of juvenile largemouth bass. In addition, using high‐fat diets, TRO diet improved the antioxidant capacity and immune ability of juvenile largemouth bass, thereby promoting the overall health of fish. This study provides novel insights for fish feed formulation optimization from the perspective of genetically modified feed ingredients, and high‐quality aquatic products for human consumption.