Abstract: In order to verify whether the cell penetrating peptides could enhance the bactericidal action of aminoglycoside antibiotics, three representative aminoglycoside antibiotics (tobramycin, kanamycin and streptomycin) were selected to treat Escherichia coli at plateau phase. Under the specific antibiotic treatment concentration and treatment time, the killing effect of Escherichia coli at plateau phase was compared between groups, and the differences of experimental results was analyzed by using the ANOVA algorithm. Then, the NPN staining method was used to explore the mechanism of cell penetrating peptides enhancing the bactericidal action of aminoglycoside antibiotics. The results showed that the pretreatment with cell penetrating peptides could extremely significantly enhance tobramycin, kanamycin and streptomycin to kill Escherichia coli at plateau phase, and the cell penetrating peptides could slightly damage the cell outer membrane of Escherichia coli.