This paper describes near infrared (NIR)-triggered on-demand adeno-associated virus (AAV) release from alginate hydrogel microbeads with heat transducer for gene therapy. As an AAV carrier, calcium alginate (Ca-Alg.) hydrogel microbeads were fabricated by injecting mixed pre-gel solution composed of sodium-alginate (Na-Alg.), AAV, and Fe3O4 particles (FPs: heat transducer) to CaCl2 solution to form ionically cross-linked instantly with a centrifuge-based microdroplet shooting device. The FPs generate heat due to localized surface plasmon resonance under NIR. By irradiating NIR to the microbeads, we confirmed the release of AAV and the gene transfection to HeLa cells. Our proposed NIR-triggered AAV release system would be an effective and safe controlled release system for gene therapy.