High-yield antibiotics producing strain, preparation method and use thereof are provided in the present invention. The high-yield strain is a mutagenized strain derived from Colephoma empetri, and deposited in CGMCC with the accession number of CGMCC 4129. The preparation method comprises the following steps: (a) mixing a seed liquid of Colephoma empetri of Accession No. FERM BP-2635 with nitrosoguanidine to obtain a mixture a; (b) mixing said mixture a with a wall-breaking enzyme to obtain protoplasts; (c) regenerating said protoplasts to obtain single colonies; and (d) culturing said single colonies to obtain said mutagenized strain. The obtained strain has stable genetic and producing property, produces little impurities in fermentation, and is suitable for industrialization.