Objective To study the rule of callus culture and baicalin synthesis in Scutellaria baicalensis. Methods Callus was induced by plant cell culture technology and the content of baicalin was determind by HPLC. Results The optimal culture medium on the growth of callus and the synthesis of baicalin in S. baicalensis is: MS culture medium, 60 mmol/L (NH_4+∶NO_3-=1∶1), 0.5—1.5 mmol/L KH_2PO_4, 80 g/L sucrose, 0.3 mg/L IAA, 2 mg/L 6-BA, and 200 mg/L peptone. When it was cultured for 40 d, the total biomass reached 28.7 g/L and the content of baicalin was 167.4 mg/g, which was much higher than that of wild S. baicalensis. Conclusion The growth of S. baicalensis callus and the accumulation of baicalin are not underway simultaneously; the callus grows first and then its secondary metabolic products synthesize. It is obvious for sucrose to regulate the baicalin synthesis. When the concentration of sucrose is less than 3%, it could only promote the callus growth; when between 3% and 8%, it could greatly increase not only the callus growth but also the baicalin synthesis, when 8%, both of them arrive to the maximum content.