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Fe3O4 nanoparticles (NPs) are widely used in the construction of drug and gene delivery vectors because of their particular physicochemical properties. Surface modification can not only reduce the cytotoxicity of Fe3O4, but also further improve the biocompatibility and delivery efficiency. In this work, firstly, polydopamine (PDA)-coated Fe3O4 NPs (named Fe3O4@PDA) were prepared by using the self-polymerization characteristics of dopamine in alkaline environment. Then, polyamidoamine (PAMAM) was modified by the Michael addition reaction to prepare water-soluble core—shell magnetic NPs of Fe3O4@PDA@PAMAM, and its potential as gene vector was further evaluated. The results revealed that Fe3O4@PDA@PAMAM had the ability to condense and protect DNA, and showed lower cytotoxicity, higher cell uptake and transfection efficiency than those of PAMAM. It has the potential to become a magnetic targeted gene vector for further study. [ABSTRACT FROM AUTHOR]