Taking the potency of phytolactin as a characteristic index, the culture conditions for bacteriocin production by Lactobacillus plantarum M1-NTG300 were optimized. Through response surface analysis combined with actual values, the optimal components of the culture medium were determined to include 3.5% glucose concentration, 3.5% peptone, 0.35% Tween 80, and 0.20% magnesium chloride. Under the control of these culture conditions, The titer of bacteriocin, produced by L. plantarum M1-NTG300 was 1 269.4 IU/mL, which was increased by 19.8%.