In this paper, He-Na laser irradiation and antibiotics resistance selection method were used to screen daptomycin high-producing strains.He-Na laser irradiation was applied to screen daptomycin high-producing strain from original strain of Streptomyces roseosporus DA15-2.The optimum mutagenesis dose was irradiation power 20 mW and irradiation time for 20 min, which was determined based on survival rate and pos. mutation rate.The mutant strains on the plates of combined antibiotics resistance of daptomycin and Streptomyces at different concentrations were screened and a mutant strain DA15-23 was obtained, whose yield of daptomycin was 100 mg/L, which increased 75.8% compared to that of the parental strain.It was demonstrated that laser irradiation and antibiotics resistance selection combined method provided a fast and effective way of screening Streptomyces roseosporus.