Abstract: To improve the β-glucosidase production, iterative ARTP was compounded with UV to breed S. cerevisiae G13 and G21 strains mutants with increased β-glucosidase production. After one round of ARTP and two rounds of UV mutagenesis, two strains of high-yielding β-glucosidase mutants, Dea-G13D1Z2 and Dma-G21D1Z2, were selected which showed increased β-glucosidase by shaking flask fermentation.After 5 consecutive generations of fermentation production tests, the enzyme production levels were stable at 240.93 and 173.38 U/mL, respectively, which were 4.61 times and 3.59 times the enzyme activities of the starting strains G13 and G21.Iterative ARTP-UV is a potentially efficient method for microbial mutation breeding and can be effectively applied to microbial breeding.