80 yeast strains were isolated from orange juice. Through the Duchenne tubule screening, alcohol production capacity screening, SO2 tolerance screening, pH tolerance screening, and GC-MS flavor screening, the optimal yeast numbered as 20 was obtained finally. The result from molecular biological identification showed that this strain belonged to the genus Saccharomyces cerevisiae and could be used for food fermentation. Compared with wine yeast D254, the total alcohol content of orange wine produced by yeast 20 increased by 50.49%, total ester content increased by 24.85%, terpenes content increased by 31.67%, acid content increased by 44.71%, phenolic compounds content increased by 59.30%.