Production of oleanane-type sapogenin in transgenic rice via expression of β-amyrin synthase gene from Panax japonicus C. A. Mey

Zhiwei Huang, Juncheng Lin, Zuxin Cheng, Ming Xu, Mingshu Guo, Xinying Huang, Zhijian Yang, Jingui Zheng

Panax japonicus C. A. Mey. is a rare traditional Chinese herbal medicine that uses ginsenosides as its main active ingredient. Rice does not produce ginsenosides because it lacks a key rate-limiting enzyme (β-amyrin synthase, βAS); however, it produces a secondary metabolite, 2,3-oxidosqualene, which is a precursor for ginsenoside biosynthesis. In the present study, the P. japonicus βAS gene was transformed into the rice cultivar 'Taijing 9' using an Agrobacterium-mediated approach, resulting in 68 rice transgenic plants of the T0 generation. Transfer-DNA (T-DNA) insertion sites in homozygous lines of the T2 generation were determined by using high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR) and were found to vary among the tested lines. Approximately 1-2 copies of the βAS gene were detected in transgenic rice plants. Real-time PCR and Western blotting analyses showed that the transformed βAS gene could be overexpressed and β-amyrin synthase could be expressed in rice. HPLC analysis showed that the concentration of oleanane-type sapogenin oleanolic acid in transgenic rice was 8.3-11.5 mg/100 g dw. The current study is the first report on the transformation of P. japonicus βAS gene into rice. We have successfully produced a new rice germplasm, "ginseng rice", which produces oleanane-type sapogenin.