High-intensity breeding of broilers has increased meat yield while reducing meat quality. Properly enhancing intramuscular fat (IMF) content is an effective strategy to improve chicken meat quality and boost consumer preference. Given that the role of phosphatidylethanolamine (PE) in chicken IMF metabolism remains unclear, we investigated the effects of PE on the meat quality, as well as the differentiation and gene expression regulation of chicken intramuscular adipocytes (IMAs). PE supplementation significantly increased the a* value of meat color and reduced shear force (P < 0.05); however, it did not exert a significant effect on the pH value 45 min post-slaughter (P > 0.05). After treating intramuscular adipocytes IMAs with 25, 50, and 100 mu M PE, 100 mu M PE supplement markedly enhanced lipid deposition and the expression of genes related to adipogenic differentiation. Differentially expressed genes (DEGs) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Set Enrichment Analysis (GESA) were used to identify key genes involved in PE regulation of chicken IMA deposition. A total of 1,182 DEGs were identified, with genes S1PR3, FABP4, PLIN2, APOA1, PPARG and CD36 recognized as hub genes associated with the triglycerides (TG) content of IMAs. PPAR signaling pathway, terpenoid backbone biosynthesis, cytokine-cytokine receptor interaction, and neuroactive ligand-receptor interaction pathway were significantly enriched in PE group. This study reveals the role of PE in chicken IMAs differentiation and lipid deposition, providing a theoretical foundation for further research into the impact of PE on IMF accumulation in broiler chickens.