SIMPLE ONE-MEDIUM FORMULATION REGENERATION OF FINGERROOT [BOESENBERGIA ROTUNDA (L.) MANSF. KULTURPFL.] VIA SOMATIC EMBRYOGENESIS S. K. TAN 1 *, R. PIPPEN 2 , R. YUSOF 3 , H. IBRAHIM 4 , N. RAHMAN 5 , AND N. KHALID 4 1 Sunway University College, 5 Jalan Kolej, Bandar Sunway, PJ 46150 Selangor, Malaysia 2 Western Michigan University, 1903 West Michigan Ave, Kalamazoo, MI 49008-5211 3 Biochemistry Department, Medical Faculty, University Malaya, 50603 Kuala Lumpur, Malaysia 4 Institute of Biological Sciences, Science Faculty, University Malaya, 50603 Kuala Lumpur, Malaysia 5 Chemistry Department, Science Faculty, University Malaya, 50603 Kuala Lumpur, Malaysia (Received 4 May 2004; accepted 30 May 2005; editor M. A. O’Connell) Summary Most published protocols necessitate different media formulations for multistep somatic embryogenesis. This study aims to establish a simple but effective formulation for the regeneration of plantlets of the pharmaceutically active Boesenbergia rotunda (L.) Mansf. Kulturpfl, formerly Boesenbergia/Kaempferia pandurata (Schult), to ensure a superior and consistent supply of materials for commercialization purposes. In this study, a single-medium formulation of Murashige and Skoog (MS) supplemented with 13.54 mM 2,4-dichlorophenoxyacetic acid (2,4-D) was found to be the only medium out of eight formulations to promote the complete somatic embryogenesis process for the culture of B. rotunda (L.). Callus cultures were initiated from a total of 280 explants of rhizome meristem. The percentage of cultures forming embryogenic callus was 23.3 ^ 4.3% on this MS medium augmented by 13.54 mM 2,4-D. The best plantlet regeneration rate was attained from the first subcultured callus with a mean of 6.6 ^ 0.1 plantlets per 1 cm diameter aggregate of callus. Somatic embryogenesis characteristic of monocots was evident from histological studies. The regenerated plantlets have been successfully established in soil. Key words: auxins; micropropagation; ginger; Zingiberaceae. Introduction Fingerroot, Boesenbergia rotunda (L) Mansf. Kulturpfl. (Larsen, 1996) (BR), formerly Boesenbergia or Kaempferia pandurata (Schult) is a folk medicinal plant used as an aphrodisiac and for treating dyspepsia. It is also used as a spice. Fingerroot is widely cultivated in South East Asia for its small underground rhizome. In recent years, there has been increasing interest in its rhizome as a good source of several potent bioactive flavanoids and chalcones. Fahey and Stephenson (2002) found that pinostrobin, a flavonone from fingerroot, can elevate the activity of quinone reductase (an antioxidant enzyme) by 500 000 units g 21 . BR compounds were also found to be antimutagenic (Murakami et al., 1993; Trakoontivakorn et al., 2001). Its cyclohexenyl chalcone derivatives were reported to be anti-inflammatory (Tuchinda et al., 2002) and Yun et al. (2003) reported that this anti-inflammatory property of one of these cyclohexenyl derivatives, panduratin A, was due to its strong inhibition of both nitric oxide and prostaglandin production through the suppression of NF-kappaB activation. Tewtrakul et al. (2003) found that cardamonin isolated from BR exhibited appreciable anti- HIV-1 protease inhibition. In earlier studies, BR also exhibited analgesic and antipyretic effects (Pathong et al., 1989). BR extract had been reported to yield various chalcones (Trakoontivakorn et al., 2001), flavonoids (Jaipetch et al., 1983), flavones (Jaipetch et al., 1982), and essential oil (Lawrence et al., 1971; Pandji et al., 1993). His species has great pharmaceutical potential, hence the importance of establishing a simple protocol for its micropropaga- tion to ensure a superior and consistent supply of materials for commercialization purposes, especially in light of its small rhizomes. Somatic embryogenesis is an efficient method that could be adopted for micropropagation and genetic transformation. However, very few reports have been published on the somatic embryogenesis of plants belonging to the family Zingiberaceae (Malamug et al., 1991; Kackar et al., 1993; Salvi et al., 2001). Somatic embryogenesis is a regeneration process that starts with nonzygotic cells transforming into a proembryogenic mass, then bipolar somatic embryos and eventually forming plantlets. It is generally accepted that the demand for plant growth factors will vary accordingly, both in nature and concentration, as the asomatic cells differentiate into somatic embryos then into plantlets, hence the need for several changes of media supplemented with tested exogenous plant growth factors. Thus, clonal propagation via somatic embryogenesis can be rather demanding and complicated as in most established protocols. All three published works on clonal propagation of two Zingiberaceae species, tumeric (Salvi et al., 2001) and ginger (Malamug et al., 1991; Kackar et al., 1993), required a few changes *Author to whom correspondence should be addressed: Email tskiat@ academic.sunway.edu.my In Vitro Cell. Dev. Biol.—Plant 41:757–761, November – December 2005 DOI: 10.1079/IVP2005695 q 2005 Society for In Vitro Biology 1054-5476/05 $18.00+0.00 757