Grouping and characterization of putative glycosyltransferase genes from Panax ginseng Meyer

Glycosyltransferases are members of the multigene family of plants that can transfer single or multiple activated sugars to a range of plant molecules, resulting in the glycosylation of plant compounds. Although the activities of many glycosyltransferases and their products have been recognized for a long time, only in recent years were some glycosyltransferase genes identified and few have been functionally characterized in detail. Korean ginseng (Panax ginseng Meyer), belonging to Araliaceae, has been well known as a popular mysterious medicinal herb in East Asia for over 2000 years. A total of 704 glycosyltransferase unique sequences have been found from a ginseng expressed sequence tag (EST) library, and these sequences encode enzymes responsible for the secondary metabolite biosynthesis. Finally, twelve UDP glycosyltransferases (UGTs) were selected as the candidates most likely to be involved in triterpenoid synthesis. In this study, we classified the candidate P. ginseng UGTs (PgUGTs) into proper families and groups, which resulted in eight UGT families and six UGT groups. We also investigated those gene candidates encoding for glycosyltransferases by analysis of gene expression in methyl jasmonate (MeJA)-treated ginseng adventitious roots and different tissues from four-year-old ginseng using quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). For organ-specific expression, most of PgUGT transcription levels were higher in leaves and roots compared with flower buds and stems. The transcription of PgUGTs in adventitious roots treated with MeJA increased as compared with the control. PgUGT1 and PgUGT2, which belong to the UGT71 family genes expressed in MeJA-treated adventitious roots, were especially sensitive, showing 33.32 and 38.88-fold expression increases upon 24 h post-treatments, respectively

Isolation and characterization of Panax ginseng geranylgeranyl-diphosphate synthase genes responding to drought stress

Geranylgeranyl-diphosphate synthases (GGDPS) catalyze branch point enzymatic reactions producing isoprenoid-derived products which are necessary for plant growth and responses to a wide range of biotic and abiotic stresses. In our study, full length geranylgeranyl-diphosphate synthase 1 (PgGGDPS1) and 2 (PgGGDPS2) cDNA were isolated and characterized from the flower of Panax ginseng and 4-year old P. ginseng cv. Gumpoong. The cDNA had open reading frame of 1032 and 1116 bp with a deduced amino acid sequence of 343 and 371 residues for GGDPS1 and GGDPS2, respectively. The calculated molecular mass of GGDPS1 and GGDPS2 were approximately 37.66 and 40.21 kDa with a predicated isoelectric point of 5.32 and 6.23 and predicted localization of plastid. A GenBank Blast X search revealed that the deduced amino acid of PgGGDPS1 shared a high degree of homology with GGDPS from Panax notoginseng. The transcription pattern of GGDPS genes was different at various developmental stages. Both GGDPS genes were highly expressed in aerial parts of the plant, especially in rapidly growing tissues such as 4-year old flower and stem tissues. Transcript level of PgGGDPS1 was differentially induced in ginseng not only during Pseudomonas syringae pv tomato infection but also after exposure to abiotic stresses. Our results suggested that the induction of GGDPS genes specifically PgGGDPS1 by drought stress may affect chlorophyll levels, intracellular GA content and accumulation of carotenoids as the precursor for higher production of ABA and possibly stomatal closure as the barrier for water loss

Plant Regeneration from Anther Culture of Panax ginseng

Abstract -The research concerned of the regeneration of plants from embryos obtained from anther cultures of ginseng (Panax ginseng C. A. Meyer). The aim was to determine the influence of the regeneration medium on the efficiency of the regeneration process. We conducted to determine the optimum conditions such as cold pretreatment, plant growth regulators and carbon sources on anther culture of P. ginseng. Highest callus formation rate was obtained when flower buds pretreated at 4℃ for 1 day. Among the treated growth regulators with various degrees of concentration in Murashige and Skoog`s (MS) medium, 4.53 μM of 2.4-dichlorophenoxyacetic acid and 4.44 μM of 6-benzylaminopurine gives the most responsive callus with the frequency of 73.89% and 129.53 g of fresh weight. When we used 3-9% of sucrose and maltose among the different kinds and various concentrations of carbohydrates, callus was formed highest 67.29% in the medium with 3% of sucrose. Shoots induced from callus supplemented with 28.9 μM of gibberellic acid and rooted in Gamborg`s B5 medium supplemented with 14.7 μM of indole-3-butyric acid

Molecular characterization of lipoxygenase genes and their expression analysis against biotic and abiotic stresses in Panax ginseng

Lipoxygenase (LOX) belongs to a family of non-heme-iron-containing fatty acid dioxygenases that are widely distributed in plants and animals. LOX involved in the synthesis of jasmonic acid and six-carbon (C6) volatiles which is necessary for plant growth and responses to a wide range of biotic and abiotic stresses. We have isolated and characterized LOX cDNA clones from Panax ginseng Meyer. From their deduced amino acid sequences, two diverse classes of 9-LOX (LOX1, LOX2, and LOX3) and 13-LOX (LOX4, LOX5) are defined in P. ginseng. A GenBank Blast X search revealed that the deduced amino acid of PgLOXs share a high degree of homology with LOX from other plants and mammals especially in three distinct motifs; motif1 harboring iron binding regions, motif2 and motif3. Chloroplast localization was predicted for PgLOX5. PgLOXs displayed organ-specific expression, highly expressed in aerial parts of the plant such as 3-year old flower, stem and leaf tissues. PgLOXs mRNAs were elevated strongly by bacterial infection. Expression of PgLOXs was differentially induced in ginseng not only by mechanical damage and methyl jasmonate but also after exposure to withholding water. Ginseng 13-LOXs positively respond to wounding that may involve in production of C6 volatiles and jasmonic acid at the wounded sites. However, the higher expression of PgLOX3 by water deficit and 82 % of the nucleotide sequence identity with the EST from severe drought-stressed leaves of Populus (CU229089.1) at +6371 bp downstream of PgLOX3 genomic DNA structure can suggest drought tolerance role for PgLOX3. Ginseng LOX genes have different expression pattern which may suggest different specific function against various environmental stresses

Overexpression of Panax ginseng sesquiterpene synthase gene confers tolerance against Pseudomonas syringae pv. tomato in Arabidopsis thaliana

Sesquiterpenes are an abundant group belonging to the terpenoid family, with a C15 structure comprise of three isoprene units. Many sesquiterpenes are volatile compounds and it act as chemical messenger in plant signalling, particularly in the defense mechanism against biotic and abiotic stresses. Panax ginseng Meyer is important medicinal herbs with various reported pharmacological efficacies in which its triterpenoid saponins, called ginsenosides, were mostly studied. However, there have been few studies on volatile sesquiterpenes compounds regulation on P. ginseng. As slow-growing perennial plant, P. ginseng received many kind of stresses during its cultivation. The pathogen attack is one of the most devastated perturbation for ginseng yield. Thus, we aimed to analyze P. ginseng STS gene (PgSTS) expressions in ginseng organs as well as mono-, sesquiterpenes contents from ginseng seedlings treated with elicitors. qRT-PCR and GC-MS analysis showed that two elicitors- salicylic acid (SA) and methyl jasmonate (MeJA) triggered PgSTS expression at different time points and significantly induced mono-, sesquiterpene yield. Overexpression of PgSTS in Arabidopsis also induced high terpene content and conferred tolerance against Pseudomonas syringae pv. tomato infection. These results suggested that PgSTS transcripts are involved in terpenoid biosynthesis in response to environmental stress mediated by MeJA and SA elicitors; thus, generate tolerance against pathogen attack

Metabolite Profiling of In Vitro Cultured and Field Grown Rhizomes of Acorus calamus from Mongolia Using GC-MS

Acorus calamus (sweet flag) is used in the traditional Chinese and Indian medicines for various ailments. Due to its extensive use in herbal medicine, natural resources from the world's forests are being depleted at an alarming rate. In the present study, an in vitro cell culture technique is being explored as an alternative to field grown A. calamus with respect to the metabolite profile, antioxidant properties, total phenol, and total flavonoid content. Gas chromatography mass spectrometry (GC-MS) was utilized to compare the metabolite profiling between methanolic extracts of in vitro and field grown rhizome tissues of A. calamus. A statistical analysis indicated an upregulation of a-selinene, which is representative of sesquiterpene ketones, and a cyclic polyol, D-pinitol, which has an insulin mimicking effect in the in vitro cultivated rhizome tissue when compared to field grown rhizomes. Significantly higher free-radical scavenging activity (IC50 69.32 mu g mL(-1)), total phenolic content (71.60 mg GAE g(-1)), and total flavonoid content (42.34 mg CE g(-1)) were observed in in vitro rhizome tissues compared with those from field grown rhizomes. These observations suggest that the in vitro cultivation of Acorus rhizomes could be exploited as an alternative to field grown A. calamus, as it is an endangered medicinal plant. The production of useful metabolites by the in vitro cultured rhizomes can be explored successfully for utilization by various food and drug industries