DEVELOPMENTAL BIOLOGY/MORPHOGENESIS Highly efficient plant regeneration via somatic embryogenesis from cell suspension cultures of Boesenbergia rotunda Sher Ming Wong & Nursafina Salim & Jennifer Ann Harikrishna & Norzulaani Khalid Received: 16 March 2013 /Accepted: 14 October 2013 /Published online: 8 November 2013 / Editor: Wagner Otoni # The Society for In Vitro Biology 2013 Abstract Boesenbergia rotunda is a perennial ginger species rich in flavonoids, flavones, and cyclohexenyl chalcone derivatives. Several of these secondary metabolites have shown promising antiviral and anticancer activities, and thus, it is important to optimize methods for robust production of clonal materials. In this study, cell suspensions were established and their growth capacities were evaluated in liquid media supplemented with varying growth regulator compositions. The highest settled cell volume of 6.1±0.3 ml with a specific growth rate of 0.0892±0.0035 was achieved by maintaining cells in Murashige and Skoog liquid media supplemented with 1.0 mg L −1 of 2,4-dichlorophenoxyacetic acid and 0.5 mg L −1 6-benzyladenine, representing a 12-fold increase in cell volume during the culture period. A somatic embryogenesis rate of 1,433.33±387.84 somatic embryos per milliliter of settled cells was achieved with an inoculation cell density of 50 μl settled cell volume and on growth regulator- free agar plates. Around half (53.5±7.9%) of the somatic embryos germinated into complete plantlets on media supplemented with 3 mg L −1 6-benzyladenine and 1 mg L −1 α-naphthaleneacetic acid. The plantlets were successfully transferred to soil and grown in the greenhouse. Phytochemical profiling via high-performance liquid chromatography analysis revealed that regenerated plantlets retained the capacity to produce and accumulate bioactive compounds. Hence, this protocol will be helpful for metabolic engineering and functional studies of genes and enzymes involved in the biosynthetic pathway of valuable compounds in B . rotunda . Keywords Boesenbergia rotunda . Fingerroot ginger . Medicinal compounds . Cell suspension culture Introduction Boesenbergia rotunda (L.) Mansf., commonly known as Chinese keys, fingerroot ginger, or Temu Kunci, belongs to the tribe of Hedychieae of the Zingiberaceae family (Tan et al. 2006). B . rotunda has a long history of traditional use as a spice and in folk medicine, and it recently has been shown to produce secondary metabolites with potential pharmaceutical value. These metabolites include cyclohexenyl chalcone derivatives, flavones, and flavanoids such as panduratin A and 4′-hydroxypanduratin A, which show anti-inflammatory activity (Tuchinda et al. 2002); pinostrobin, pinocembrin, and cardamonin, which show anti-HIV activity (Tewtrakul et al. 2003); isopanduratin A, possessing antimicrobial activity against the oral bacteria Streptococcus mutans (Hwang et al. 2004) and the spoilage bacteria Lactobacillus plantarum (Pattaratanawadee et al. 2006); and pinocembrin chalcone, an anti-mutagenic agent (Trakoontivakorn et al. 2001) that has antibacterial activity (Bremner and Meyer 1998). Tan et al. (2006) demonstrated that B . rotunda methanolic extract inhibited dengue 2 virus NS2b/3 protease and hence is a potential lead compound for drugs against dengue virus, which is a prevalent agent of disease in tropical and subtropical countries. Morikawa et al. (2008) extracted 18 compounds known to accumulate in rhizomes of B . rotunda , along with eight new compounds, including four prenylchalcones (krachaizin A and krachaizin B) and four prenylflavones (rotundaflavones). S. M. Wong : N. Salim : J. A. Harikrishna : N. Khalid Institute of Biological Sciences, Faculty of Science, Kuala Lumpur, Malaysia N. Khalid UMBio, University of Malaya, Kuala Lumpur, Malaysia J. A. Harikrishna : N. Khalid (*) Centre of Biotechnology for Agriculture Research (CEBAR), University of Malaya, Kuala Lumpur, Malaysia e-mail: lani@um.edu.my In Vitro Cell.Dev.Biol.—Plant (2013) 49:665–673 DOI 10.1007/s11627-013-9570-4