The optimization of fermentation medium is important for improving microbial production or quality in fermentation industry. Experimental design and optimization techniques play an important role for optimizing medium. In this paper, we reviewed the methodologies and techniques used in optimization of microbial medium, including one-factor-at-a-time design, factorial design, uniform design, response surface optimization (RSM), artificial neural network (ANN) and genetic algorithms (GA). Meanwhile, the methodologies and techniques were analyzed and evaluated comprehensively.