Agrobacteriummediated transient gene expression has become an effective method for studying gene function and producing active proteins due to its simple operation, high reproducibility and low experimental cost. In this study, the transient expression of the reporter gene GFP in the leaves of five Chinese herbal herbs ［Glycyrrhiza uralensis Fisch., Scutellaria baicalensis Georgi, _{Astragalus membranaceus} var. mongholicus (Bge.) Hsiao, Rheum officinale Baill., Isatis tinctoria L.］ was conducted by means of leaf osmotic method, which was mediated by nonviral (vector I), viral binary vector (vector Ⅱ) and two kinds of Agrobacterium tumefacients (EHA105, LBA4404). The factors affecting the expression were analyzed in order to identify the suitable materials for the application of transient expression technology in Chinese medicinal materials. The results showed that: (1) the transient transformation efficiency of Agrobacterium tumefaciens in the leaves of five Chinese herbal medicines was affected by vectors, strains and plant species types. (2) Among the five selected Chinese herbal plants, compared with the strains without transformation vector, when the OD₆₀₀ of Agrobacterium resuspend was 0.8 and the leaves were infected for 5-7 days, the fluorescence intensity of EHA105 strain containing nonviral binary vector was the highest in Glycyrrhiza uralensis Fisch. leaves, followed by Rheum officinale Baill.. The strain LBA4404 containing the virustype binary vector had the highest fluorescence expression of GFP in the leaves of Scutellaria baicalensis Georgi. Fluorescence or almost no fluorescent protein expression was detected in the leaves of Glycyrrhiza uralensis Fisch. and Astragalus membranaceus var. mongholicus (Bge.) Hsiao membranaceus infected by different bacterial fluids containing different vectors, that is, they were not diffused to the surrounding areas. Weak fluorescence or no fluorescence intensity was detected in the leaves of Isatis tinctoria L.. The results showed that the leaves of Glycyrrhiza uralensis Fisch. and Scutellaria baicalensis Georgi could be used as materials for Agrobacteriummediated transient gene expression.