Abstract: Improving the edible quality of gluten-free cereal foods has always been a hot topic for scientists. In this study, zein-based gluten-free dough was prepared with zein, corn starch, and Artemisia sphaerocephala Krasch gum as raw materials. Using two methods of introducing lactic acid, by adjusting the ratio of lactic acid to zein in dough (L/Z ratio), the rheological properties of zein-based gluten-free dough and the microstructure of zein were investigated. The study found that increasing of L/Z ratio in dough would lead to the decrease of storage modulus (G′) and the increase of loss coefficient (tanδ) of lactic acid added group (T dough) and lactic acid pretreated zein-based dough (C dough). Different ways of lactic acid introduction would lead to different rheological properties of dough. When the L/Z ratio was constant, the G' of T dough was lower than that of C dough, and the tanδ was higher than that of C dough. Compared with C dough, T dough had more practical value. As the L/Z ratio in the dough increasing, the tensile properties of T dough increased first and then decreased, when the L/Z ratio increased from 0:10 to 2:10 (T0~T2), the stretch of the T dough gradually increased, the mass aggregation of zein in dough decreased, and connected into a network gradually. T2 had the best tensile properties, the texture of zein network in dough was uniform, and had the application potential. As T3~T5, the stretchability of the dough gradually decreased, the network of zein in dough became weaker, and the level of starch granules encapsulation decreasesd. C dough had poor stretchability and could not effectively to carry out related experiments, which might be related to the failure to build a strong and coherent protein network in the dough.