ORIGINAL PAPER Production of the dammarene sapogenin (protopanaxadiol) in transgenic tobacco plants and cultured cells by heterologous expression of PgDDS and CYP716A47 Ju-Hyeon Chun 1 • Prakash Babu Adhikari 1 • Seong-Bum Park 1 • Jung-Yeon Han 1 • Yong-Eui Choi 1 Received: 10 March 2015 / Revised: 21 April 2015 / Accepted: 6 May 2015 Ó Springer-Verlag Berlin Heidelberg 2015 Abstract Key message Protopanaxadiol (PPD) is an aglycone of dammarene-type ginsenoside and has high medicinal values. In this work, we reported the PPD production in transgenic tobacco co-overexpressing PgDDS and CYP716A47. Abstract PPD is an aglycone of ginsenosides produced by Panax species and has a wide range of pharmacological activities. PPD is synthesized via the hydroxylation of dammarenediol-II (DD) by CYP716A47 enzyme. Here, we established a PPD production system via cell suspension culture of transgenic tobacco co-overexpressing the genes for PgDDS and CYP716A47. The concentration of PPD in transgenic tobacco leaves was 2.3–5.7 lg/g dry weight (DW), depending on the transgenic line. Leaf segments were cultured on medium with various types of hormones to induce callus. Auxin treatment, particularly 2,4-D, strongly enhanced the production of DD (783.8 lgg -1 DW) and PPD (125.9 lgg -1 DW). Treatment with 2,4-D enhanced the transcription of the HMG-Co reductase (HMGR) and squalene epoxidase genes. PPD production reached 166.9 and 980.9 lgg -1 DW in a 250-ml shake flask culture and in 5-l airlift bioreactor culture, respectively. Keywords CYP716A47 Á Dammarenediol-II synthase Á Protopanaxadiol Á Sapogenin Á Transgenic tobacco Abbreviations BAR Basta DD Dammarenediol-II PPD Protopanaxadiol DDS Dammarenediol-II synthase PgDDS Panax ginseng dammarenediol-II synthase CYP716A47 Cytochrome P450 716A47 qPCR Real-time polymerase chain reaction Introduction The root of Panax ginseng is one of the most famous and widely used medicinal plant materials (Shibata 2001). The major pharmacologically active components of ginseng are dammarene-type ginsenosides, a diverse group of triter- penoid saponins. More than 30 types of ginsenosides have been identified. Dammarene-type ginsenosides are the major saponin constituents in Panax species (Shibata 2001) and are divided into two groups based on their aglycone structure: protopanaxadiol (PPD) and protopanaxatriol (PPT). Ginseng roots and their products are typically adminis- tered orally. The absorption of glycosylated ginsenosides by the gastrointestinal tract is extremely low (Tawab et al. 2003). Acid or intestinal bacteria in the gastrointestinal tract degrade ginsenosides. Deglycosylated ginsenosides are more readily absorbed into the bloodstream and func- tion as active compounds (Karikura et al. 1991). Hy- drolyzed ginsenosides or ginsenoside aglycones have Communicated by F. Sato. Electronic supplementary material The online version of this article (doi:10.1007/s00299-015-1806-9) contains supplementary material, which is available to authorized users. & Yong-Eui Choi yechoi@kangwon.ac.kr 1 Department of Forest Resources, College of Forest and Environmental Sciences, Kangwon National University, Chunchon 200-701, Republic of Korea 123 Plant Cell Rep DOI 10.1007/s00299-015-1806-9