RNAi expression vector of cycloartenol synthase gene (PjCAS) from Panax japonicus was constructed by Gateway technology in this study, and Agrobacteriummediated transformation was used to realize RNAi of PjCAS in P. japonicus cells successfully. Realtime quantitative PCR was used to analyze the expression of key enzyme genes involved in the biosynthetic pathway of P. japonicus saponins (PJS), the changes of saponins and phytosterols in transgenic cells were also determined, and the regulation effect of PjCAS on the synthesis of PJS was discussed. The results showed that: (1) the RNAi fragment of PjCAS was amplified, and the PjCAS RNAi vector pHellsgatePjCAS was successfully constructed. (2) Six positive transgenic cell lines with PjCAS interferance were obtained by Agrobacterium transformation. (3) Compared with the ordinary cell line, the expression of PjCAS in transgenic cell lines was approximately decreased by 85%, and the highest expression levels of key enzyme genes PjDS and PjAS directly related to saponins synthesis were upregulated by 90% and 150%, respectively. (4) The contents of six monomeric saponin in the transgenic cell lines were significantly higher than that of control, among them, the average contents of dammaranetype monomeric saponin Re, Rb1, Rd and oleananetype monomeric saponin R0, IV, IVa were higher than that of the normal P. japonicus cells, increased by 28%, 49%, 40%, 36%, 59%, and 50%, respectively. The results indicated that the change of PJS content is indirectly regulated by PjCAS gene. (5) The phytosterol content in six transgenic cell lines decreased by 53%-73% than that in control group. Studies have found that silencing PjCAS gene can significantly upregulate the expression of key enzyme genes PjDS and PjAS related to the synthesis of PJS, and increase the content of monomeric saponin in the cell lines with PjCAS, thus promote the significant increase in the synthesis of PJS. The results prove that the metabolic flux of the branch of phytosterol synthesis could be decreased by inhibiting the expression of PjCAS, the key gene in the biosynthetic pathway of phytosterols. As a result of it, more metabolic flux flowed towards the synthesis of PJS and the biosynthsis of PJS was promoted.