This study aimed to explore the optimal culture conditions for transcriptional expression of leukocidin ED (lukED) of Staphylococcus aureus (S. aureus). The transcription level of lukE in S. aureus strain Newman was detected at different growth phases (early-exponential, middle-exponential, late-exponential and stationary phases) in TSB, BHI, YCP, MHB and LB media by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Based on the medium mediating the lowest transcription level of lukE, the different compositions of medium producing the highest mRNA level of lukE was added to discover the substances promoting lukED expression. The findings showed that lukE mRNA level was highest at the late-exponential phase in YCP medium, in which yeast extract, casamino acids and sodium pyruvate could all strikingly increase the expression of lukE, with the first two being better.