Identification and characterization of class I chitinase in Panax ginseng C. A. Meyer.

The role of plant chitinases in protecting plants against a variety of fungal pathogens is well established. In the present study, a cDNA clone containing a class I chitinase (Chi-1) gene, designated as PgChi-1, has been isolated from the oriental medicinal plant Panax ginseng. PgChi-1 is predicted to encode a protein of 34.9 kDa consisting of 323 amino acid residues. PgChi-1 was found to be expressed constitutively in all of the studied organs of ginseng plant. Under various abiotic stress treatments including Cu, H2O2, mannitol, SA, JA, and NaCl, the expression of PgChi-1 in plantlets and hairy roots increased significantly compared to the control. When different parts of root were analyzed, maximum level was observed in taproot. In addition, levels of PgChi-1 expression were compared between healthy root and fungal, bacterial, and nematode infected root. Significant increase of PgChi-1 was noticed in pathogen infected roots than healthy roots. This study revealed that PgChi-1 may protect the P. ginseng under both biotic and abiotic stress conditions.

Inhibitory effects of the transgenic Panax ginsengs on phorbol ester plus A23187-induced IL-6 production and cyclooxygenase-2 via suppression of NF-κB and MAPKs in HMC-1.

Our previous studies have that demonstrated the overexpression of the squalene synthase gene enhances the biosynthesis of triterpene and phytosterol in Panax ginseng. The total ginsenoside contents in adventitious roots of transgenic P. ginseng were about 1.6-3-fold higher than those in the wild-type. In the present work, we have evaluated the anti-inflammatory effects of two types of transgenic P. ginseng (BS and SS) and the wild-type P. ginseng (GS) in a stimulated human mast cell line 1 (HMC-1). GS, BS, and SS inhibited not only the production of interleukin 6 (IL-6), but also the expression of cyclooxygenase-2 in phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore A23187 (PMACI)-stimulated HMC-1. Additionally, GS, BS, and SS suppressed the expression of the nuclear transcription factor κB and mitogen-activated protein kinases induced by PMACI. The anti-inflammatory effects of BS and SS were higher than that of GS. These results provide new insights into the pharmacological actions of transgenic P. ginseng as a potential molecule for use in therapy in mast cell-mediated inflammatory diseases.

Isolation and characterization of a Glutamate decarboxylase (GAD) gene and their differential expression in response to abiotic stresses from Panax ginseng C. A. Meyer.

Glutamate decarboxylase (GAD) catalyzes the conversion of L-glutamate to γ-aminobutyric acid (GABA). A full-length cDNA encoding GAD (designated as PgGAD) was isolated and characterized from the root of Panax ginseng C. A. Meyer. The length cDNA of PgGAD was 1881 bp and contained a 1491 bp open reading frame (ORF) encoding a glutamate decarboxylase protein of 496 amino acids, possessing a Ser-X-X-Lys active site, which belongs to the GAD group. The deduced amino acid sequence of the PgGAD was classified in the plant GAD family and has 76-85% high similarity with other plants as like petunia, Arabidopsis, tomato. Secondary structure of PgGAD was predicted by using SOPMA software program. Southern blot analysis of genomic DNA suggests that, there is more than one copy of the PgGAD gene. The organ specific gene expression pattern also studied in P. ginseng seedlings, in which the stem showed elevated expression than root, leaf, bud and rhizomes. Along with this, we also confirmed the gene expression of PgGAD under various abiotic stresses like temperature stress, osmotic stress, anoxia, oxidative stress, and mechanical damage. Temporal analysis of gene expression except exposure of oxidative stress revealed an enhanced expression after each stresses. The enzyme activity of PgGAD was stimulated to 2-fold under cold stress.

Isolation of S-adenosyl-L-methionine synthetase gene from Panax ginseng C.A. meyer and analysis of its response to abiotic stresses.

A cDNA clone containing a S-adenosyl-L-methionine synthetase (SAMS) gene, named as PgSAM, was isolated from a commercial medicinal plant Panax ginseng. PgSAM is predicted to encode a precursor protein of 307 amino acid residues, and its sequence shares high homology with a number of other plant SAMS. PgSAM is expressed at different levels in various organs of ginseng. The expression of PgSAM in adventitious roots and hairy roots of P. ginseng were analyzed using reverse transcriptase (RT)-PCR and real-time PCR under various abiotic stresses. Salt, salicylic acid, abscisic acid and chilling stresses induced PgSAM significantly at different time points within 2-72 h post-treatment. This study revealed that PgSAM may help to protect the plants against various abiotic stresses.

Isolation and characterization of a novel short-chain alcohol dehydrogenase gene from Panax ginseng.

The cDNA of alcohol dehydrogenase (PgADH) was isolated and characterized from the leaf of Panax ginseng. The cDNA had an open reading frame of 801 bp and a deduced amino acid sequence of 266 residues. The calculated molecular mass of the mature protein is approximately 29 kDa with a predicated isoelectric point of 6.84. Homology analysis revealed that the deduced amino acid of PgADH shares a high degree of homology with the short-chain ADH proteins of other plants. Genomic DNA hybridization analysis indicated that PgADH represents a multi-gene family. The expression of PgADH under various environmental stresses was analyzed at different time points using real-time PCR. ABA, SA and especially JA (80-fold) significantly induced PgADH expression within 24 h of treatment. The positive responses of PgADH to abiotic stimuli suggest that ginseng ADH may protect against hormone-related environmental stresses.