Provided are silica nanoparticles doped with dyes having a negative charge by using cationic polyelectrolyte as templet and preparing method thereof. The nanoparticles have a core-shell structure, inner core comprises cationic polyelectrolyte aggregates, in which fluorescent dye having a negative charge and SiO2 are doped, and optional additive having a negative charge, the shell is consisted of SiO2, wherein the sum of negative charge carried by the additives and the fluorescent dye is less than that of positive charge carried by the polyelectrolyte aggregates. The preparing method comprises the following steps: preparing complex solution of polyelectrolyte and fluorescent dye; prehydrolysis of organosilicon source; coating with organosilicon source. The method can be applied to dope silica with various fluorescent dyes having a negative charge. Various dyes can be doped at the same time, the doping amount, and particle diameter and shape of the product can be controlled by the method. The nanoparticles are spherical, the particle diameter thereof is uniform, and colloid formed therefrom is stable.