Natural killer (NK) cells are considered immunotherapeutic effectors in the fight against cancers. However, genetic materials' transfer remains a serious problem due to the low efficiency of transfection in NK cells. Here, we report that magnetic nanoparticles (MNPs) with a cationic polymer layer can genetically engineer NK cells and follow them in vivo through magnetic resonance (MR) and fluorescent imaging. These nanoparticles were dispersible in water, PBS, and cell culture medium. Also, MNPs quickly formed gene-complexes due to immobilized cationic polymer by a polydopamine. The MNPs-mediated transfection preserved the functional phenotype of NK cells. Interestingly, these nanoparticles produced EGFR-chimeric antigen receptor (CAR) NK cells, which had a significant cytotoxicity in breast cancer cells. The EGFR-CAR NK cells modified by MNPs also encompassed an anti-tumor effect in xenograft mice model and the high accumulation into the tumor. The MNPs with a cationic polymer layer could be an innovative platform for enhanced cancer immunotherapy.