Docosahexaenoic acid (DHA) is an essential nutrient for humans and plays a critical role in human development and health. Freshwater fish, such as the common carp (), have a certain degree of DHA biosynthesis ability and could be a supplemental source of human DHA needs. The elongase of very-long-chain fatty acid 5 (Elovl5) is an important enzyme affecting polyunsaturated fatty acid (PUFA) biosynthesis. However, the function and regulatory mechanism of the gene related to DHA synthesis in freshwater fish is not clear yet. Previous studies have found that there are two copies of the gene, and which have different functions. Our research group found significant DHA content differences among individuals in Yellow River carp ( var.), and four candidate genes were found to be related to DHA synthesis through screening. In this study, the expression level of is decreased in the high-DHA group compared to the low-DHA group, which indicated the down-regulation of in the DHA synthesis pathways of Yellow River carp. In addition, using a dual-luciferase reporter gene assay, we found that by targeting the 3'UTR region of , miR-26a-5p could regulate DHA synthesis in common carp. After CRISPR/Cas9 disruption of , the DHA content in the disrupted group was significantly higher than in the wildtype group; meanwhile, the expression level of in the disrupted group was significantly reduced compared with the wildtype group. These results suggest that may be down-regulating DHA synthesis in Yellow River carp. This study could provide useful information for future research on the genes and pathways that affect DHA synthesis.