Stems, petioles and leaves of four peony cultivars called Dafugui，Taohuafeixue，Karl Rosenfield and Sarah Bernhardt were selected to study major factors affecting the induction and regeneration of peony callus. The best peony cultivars was Dafugui and Karl Rosenfield in callus induction. Stem had an better effect than petiole. The best medium for callus induction was WPM+2.0 mg/L 6-BA +0.2 mg/L NAA +0.2 mg/L 2,4-D. It was better to sample explants in stooling stage than in leaf-expansion period. The optimum medium for callus proliferation were 1.0 mg/L NAA +2.0 mg/L 6-BA +0.1 mg/L TDZ for Dafugui and 1.0 mg/L NAA +0.5 mg/L 6-BA +0.1 mg/L TDZ for Karl Rosenfield. It was hard to induce adventitious bud，only some white bud points can be seen. The optimum medium for adventitious root induction was WPM+0.5 mg/L IBA, and the rooting rate was 50.00%.