Abstract:[Background] The commonly known poly-γ-glutamic acid (γ-PGA) producers are Bacillus subtilis, B. amyloliquefaciens, and B. licheniformis, while little is known about B. siamensis. [Objective] To study liquid fermentation conditions for γ-PGA production by B. siamensis. [Methods] B. siamensis CAU83, isolated by our laboratory, was used to produce γ-PGA by liquid fermentation. The effects of carbon sources, nitrogen sources, precursors, temperature, and pH on the synthesis of γ-PGA in shake flasks were investigated by single factor test and orthogonal design. [Results] The optimal carbon source, nitrogen source, and precursor for the synthesis of γ-PGA were 30 g/L lactose, 5 g/L yeast extract, and 60 g/L L-sodium glutamate, respectively. The optimal fermentation conditions were 37 ℃ and pH 7.0. The yield of γ-PGA increased by 260% from 8.4 g/L before optimization to 30.1 g/L after optimization. The fed-batch fermentation showed the peak yield (59.5 g/L) of γ-PGA at the time point of 60 h, with a productivity of 0.99 g/(L·h), which increased by 98% compared with the yield in shake flasks. The produced γ-PGA had a molecular weight of 3.8×106 Da and a high polymerization degree. [Conclusion] The optimal fermentation conditions for B. siamensis CAU83 producing γ-PGA determined in this study provide a basis for the industrial production and application of this strain.