This study demonstrated the expression of mutant phytase was found to be optimal at a temperature of 30C. The 5 % of extra yeast extract provided in the induction phase increased phytase yield (23,494.3 U g1). However, the over-expression in recombinant E. coli was essentially affected by the presence of acetic acid (>2.0 g L1). The comparison of fermentation processes concluded that the specific activity of phytase can be significantly improved from 226.74 to 498.47 U mg1. A two-stage fermentation strategy can be employed to achieve high production of phytase. This prominence could be an advantage for the downstream processing of phytase.