Abstract: In order to establish a high frequency regeneration system of Petunia hybrida Meilin, the effects of different ratios of growth regulators, different parts of leaves and different forms of callus on the regeneration of adventitious buds were studied. The results showed that 6-BA, NAA and their ratios had significant effects on the regeneration rate of adventitious buds of Petunia hybrida. Among which, 0.3 mg·L^-1 NAA significantly promoted the regeneration of adventitious buds of Meilin. When the concentration of 6-BA and NAA respectively exceeded 1.0 mg·L^-1 and 0.5 mg·L^-1, the occurrence of vitrification was stimulated. Different parts of leaves significantly affected the regeneration of adventitious buds. The bud differentiation rate from high to low was petiole, with main vein, and without main vein. The ability and quality of bud differentiation were determined by the shape of callus. The direct differentiation of cluster buds from petioles was an excellent explant for the regeneration of Meilin, with the differentiation rate of 98.3%. The optimal medium for the induction of bud was MS+6-BA 1.0 mg·L^-1+NAA 0.3 mg·L^-1, the adventitious buds were produced without vitrification and the differentiation rate was high, reaching 68.3%. The optimal medium for the induction of leaf callus was MS+6-BA 1.0 mg·L^-1+NAA 0.3 mg·L^-1, and the differentiation rate of adventitious bud was 68.3% without vitrification. The best rooting medium was 1/2 MS medium, in which the rooting rate could reach 100.0%. The optimal medium for the multiplication culture was MS+6-BA 0.5 mg·L^-1+NAA 0.1 mg·L^-1, and the cluster buds were abundant and uniform in size.