The present invention relates to a low serum efficiency mycoplasma gallisepticam attenuated strain culture medium and a preparation method thereof, and belongs to the technical field of veterinary biology. The culture medium comprises: (1) a base culture medium; and (2) an auxiliary culture medium, wherein the auxiliary culture medium mainly comprises MEM, yeast extract powder, tryptone, glucose, an inorganic salt, and the like, and growth, high titer and stability of the semi-finished product can be ensured with the auxiliary culture medium. According to the present invention, a titer of the semi-finished product bacterial liquid prepared by using the preparation method is up to 10<11> CCU/ml; and the culture medium adopts reduced pig serum to culture mycoplasma gallisepticam so as to reduce allergic stress reactions on chicken by heterologous pig serum, consider animal biosafety, improve antigen titer, and reduce production cost.