A simple fed-batch process with pre-determined exponential feeding strategy for high-cell-density cultivation of recombinant E. coli BL21 (DE3) in defined medium was developed. In this feeding method glucose and glycerol were used as the sole sources of carbon and energy to increase the cell density exponentially at controlled specific growth rates, which do not cause the accumulation of acetate. Thus, sophisticated feedback control or extra equipment to prevent the accumulation of toxic level of acetate is not necessary. The final cell densities of 100 and 118 gl-1 of dry cell mass for recombinant E. coli producing human interferon-g (hIFN-g) were obtained by using glucose and glycerol, respectively. The concentration of acetate was always maintained below toxic level. The specific yield of hIFN-g with glucose and glycerol was 93 and 92 mgg-1 of dry cell mass, and the overall productivity of hIFN-g was 0.16 and 0.14 gl-1h-1 for these two carbon sources, respectively.Keywords: Human interferon-gamma, fed-batch cultivation, recombinant E. coli, specific growth rate, glucose, glycerol