In order to improve the production of acarbose,protoplasts of an acarbose-producing strain Actinoplanes utahensis ZJB-08196 were prepared and treated with ultraviolet irradiation.The effects of factors including glycine concentration,mycelium age,lysozyme concentration,treating time,temperature and mycelial concentration on protoplast formation and regeneration were investigated.Moreover,the effect of the addition of L-proline,bovine serum albumin(BSA),polyvinylpyrrolidone(PVP) into regeneration medium on protoplast regeneration was further examined.The results indicated that protoplasts preparation and regeneration were achieved with following method.The 56 h-old seeds were inoculated into mycelia medium with 0.4% glycine and cultured for 22 h.Then the mycelia were collected and treated by 10 mg/mL lysozyme at 35 ℃ for 4 h and then spread on the R2YE-agar(regeneration medium supplemented with L-proline 4 g/L and PVP 0.2 g/L) plates.The maximum rate of protoplast regeneration reached 15.14%.After ultraviolet irradiation,a high-yield mutant UN-52 was obtained.Acarbose production of 5 165.2 mg/L with mutant UN-52 was obtained,increased by 12% compared with the original strain.