Down-regulation of β-amyrin synthase gene expression by RNA interference led to reduced levels of β-amyrin and oleanane-type ginsenoside as well as up-regulation of dammarane-type ginsenoside level. In the biosynthetic pathway of ginsenosides, β-amyrin synthase catalyzes the reaction from oxidosqualene to β-amyrin, the proposed aglycone of oleanane-type saponins. Here, RNAi was employed to evaluate the role of this gene in ginsenoside biosynthesis of Panax ginseng hairy roots. The results showed that RNAi-mediated down-regulation of this gene led to reduced levels of β-amyrin and oleanane-type ginsenoside Ro as well as increased level of total ginsenosides, indicating an important role of this gene in biosynthesis of ginsenoside. Expression of key genes involved in dammarane-type ginsenoside including genes of dammarenediol synthase and protopanaxadiol and protopanaxatriol synthases were up-regulated in RNAi lines. While expression of squalene synthase genes was not significantly changed, β-amyrin oxidase gene was down-regulated. This work will be helpful for further understanding ginsenoside biosynthesis pathway.