The invention discloses a preparation method of silicon-coated nano magnetic spheres. Magnetic spheres adsorbing silicon spheres are obtained through preparing a silicon sphere solution containing silicon dioxide nanoparticles, preparing the magnetic spheres and stirring and mixing the silicon sphere solution and the magnetic spheres; the magnetic spheres adsorbing the silicon spheres are transferred into an alcohol-water system and the silicon spheres form silicon layers on the surfaces of the magnetic spheres by using a TEOS sol-gel method; and the product silicon-coated nano magnetic spheres are obtained through washing after magnetic separation. The preparation method of the silicon-coated nano magnetic spheres is simple, safe, low in raw material cost and suitable for industrial large-scale production; the prepared nano magnetic spheres are good in dispersity and high in magnetic responsiveness; and the monodisperse silicon-coated nano magnetic spheres have a wide application prospect in the fields of bioseparation, environment, catalysis, solid-phase extraction and the like.