The effect of the quality and quantity of the inoculum on the biosynthesis of tobramycin was studied. The medium with corn steep liquor and glycerol was found to be the optimal seed medium for the tobramycin-producing organism. It is required that the fermentation medium be supplemented with 2 per cent of the vegetative inoculum at the stationary growth phase or with an increased amount of the seed material at the exponential growth phase or at the early stage of the culture dying. Subculturing of the inoculum decreases the level of tobramycin production at the fermentation stage: when the culture of the 5th--6th generation is used, the activity of the fermentation broth decreases by 30--50 per cent.