The effects of phosphate on the fermentation of L-isoleucine was studied with L-isoleucine producing strain E.coli TRFC.The effects of initial phosphate concentration in the fermentation process on biomass,the specific growth rate,yield of L-isoleucine,concentration of byproduct acetic acid and ammonium were studied by carrying out fed-batch fermentation of E.coli TRFC in 30-Liter fermentor.The results showed that maintaining the initial phosphate at the concentration of 2g/L and adding 2g/L phosphate into the broth in the late period of fermentation could effectively slow down bacterial recession.The biomass and production of L-isoleucine increased by 24.5% and 12.7%,while the amount of by-product acetic acid decreased significantly.