Chang-Yeon Yu

Intraspecific relationship analysis by DNA markers and in vitro cytotoxic and antioxidant activity in Eleutherococcus senticosus

To analyse genetic relationships and intraspecific variation within Eleutherococcus senticosus, the polymerase chain reaction (PCR) was performed on total genomic DNAs of 10 Eleutherococcus collections. Ten primers were used for amplification, yielding 106 bands, of which 87 were polymorphic. The genetic diversity and genetic distance among 10 collections of Eleutherococcus species were used to describe the dendrogram showing the phylogenic relationship. The 10 collections were classified into two groups (groups I and II) at a similarity coefficient of 0.50. Group I included E. senticosus from Bukhaedo (Japan), E. sessliliflorus from Youngwal (Korea), E. seoulense and E. chiisanesis, while group II included several internal and Russian collections. The range of polymorphism was from 66.7 to 90.9% in the 87 amplified polymorphic DNA fragments. The similarity value of all collections ranged from 0.41 to 0.92, and the average genetic distance was 0.61. Thus, RAPD analysis was useful in determining genetic relatedness among collections and in identifying different genotypes of E. senticosus and other Eleutherococcus species. Also, the biological activity on DPPH radical scavenging, antilipid peroxidation in rat liver microsomes and cytotoxic sulforhodamine B (SRB) assay was evaluated using root extracts of E. senticosus, Odaesan, Korea. Ethyl acetate and n-butanol fractionation revealed strong antioxidant against scavenging on DPPH free radical and also ethyl acetate fractionation exhibited high antilipid peroxidative activities. In the cytotoxic effects were evaluated on seven human cancer cell lines, the values of 50% growth inhibition (GI(50)) were mostly below 30 microg/ml for crude extracts to be considered as significantly active.

Overexpression of the γ-TMT gene in Codonopsis lanceolata.

A cDNA-encoding γ-tocopherol methyltransferase (γ-TMT) from Arabidopsis thaliana was overexpressed in deoduck (Codonopsis lanceolata L.) to improve the tocopherol composition. Deoduck (T2) containing the γ-TMT transgene was produced by Agrobacterium-mediated transformation. Transgene expression was confirmed by polymerase chain reaction and RNA gel blot analysis. The transgenic plants produced more leaves than control plants. In addition, the transgenic plants showed higher levels of the CSOD, CTRX, CAPX, CNADP+-IDCH, and CSO transcripts and higher SOD-like activity compared with the control plants.

Morphological changes and increase of resistance to oxidative stress by overexpression of the LebZIP2 gene in Nicotiana benthamiana

The tomato bZIP2-encoding gene was inserted into the Nicotiana benthamiana genome using Agrobacterium-mediated transformation to characterize resistance to oxidative stress and two herbicides, glyphosate and paraquat. We produced transgenic tobacco plants using the LebZIP2 gene, which were then utilized to examine salt stress and herbicide resistance through oxidative mechanisms. Transgenic LebZIP2-overexpressing plants were examined using specific primers for selection marker genes (PCR using genomic DNA) and target genes (RT-PCR). Based on microscopic examination, we observed an increase in leaf thickness and cell number in transgenic plants. The electrolyte leakage of leaves suggested that LebZIP2-overexpressing lines were weak tolerant to NaCl stress and resistant to methyl viologen. During our analysis, transgenic lines were exposed to different herbicides. Transgenic plants showed an increased tolerance based on visual injury, as well as an increased biomass. Based on these results, the LebZIP2 gene may be involved in oxidative stress tolerance and cell development in plants.

Antifungal Gene (Rs-AFP) Introduction into Rehmannia glutinosa and Gene Expression Mediated by Agrobacterium tumefaciens

Rehmannia glutinosa is one of the most important medicinal crops in Korea. However, various plant pathogens, including Fusatium spp., cause great damage on R. glutinosa and result in enormous economic losses. This study was conducted to breed Fusarium-resistant plants by using Agrobacterium tumefaciences and AFP (anti-fungal protein) gene. The plant material used was a native accession of R. glutinosa. The PCR analysis was conducted to verify transgenicity. Based on the PCR analysis, nptII band was observed in transgenic plant genome. Southern blot and AFP protein analyses also showed the expression of this gene in transgenic plants. Expression of AFP in transgenic plants offers the possibility of developing resistance to fungal infection.

Molecular characterization and stress signaling of the Miscanthus sinensis MsCOMT gene by transient assay

The full-length 1089-bp cDNA of caffeic acid 3-O-methyltransferase (COMT) was isolated from a Miscanthus sinensis cDNA library and was found to encode a 362-amino-acid-long protein. The analysis of the deduced tomato MsCOMT amino acid sequence revealed that it shares 98, 95, 92, and 80% sequence homologies with the respective proteins in Saccharum, Sorghum, maize, and Festuca arundinacea. The transcript level of the MsCOMT gene responded only to ABA signaling as compared to other environmental stressors such as mannitol, NaCl, and methyl viologen (MV) related to drought, salt, and ROS. However, MsCOMT expression decreased following the treatments with NaCl and MV. We analyzed NbPAL (Nicotiana benthamiana phenylalanine ammonia-lyase) gene expression in the leaves of Nicotiana benthamiana plants in response to the antisense MsCOMT-AS at the period ranging from 0 to 48 h. The PAL transcript level decreased when the antisense MsCOMT gene was transiently overexpressed. PR (Pathogenesis-Related) group genes led to a decrease in MsCOMT-AS gene expression in leaves. The NbAPX (Nicotiana benthamiana ascorbate peroxidase) expression level decreased following a decrease in NbPAL and PR group gene expression.

Phamalogical effect and component of sea buckthorn (Hippophae rhamnoides L.)

Abstract Sea buckthorn ( Hippophae rhamnoides L.) is deciduous shrubs in the genus Hippophae, mainly cultivated in Europe and Asia. Sea buckthorn berries have a high vitamin C, vitamin E, carotenoids, carbohydrates, protein, organic acids, dietary minerals, triterpenoids, polyphenolic acids and amino acids. Extracts of sea buckthorn berries have anti- obesity, anti-oxidantive, anti-microbial, anti-ulcerogenic, anti-diabetic and nutritional effects. Sea buckthorn used as a traditional medicine for the treatment of cough, aid di-gestion, invigorate blood circulation and alleviate pain. Ex-tracts of sea buckthorn branches and leaves was administered to humans and animals to treat gastrointestinal distress in Mongolia. This paper briefly reviews the most relevant experimental data on the pharmacological effects and iso-lated component of sea buckthorn. And, we also describe the importance of sea buckthorn as the environmental-friendly crops. Keywords anti-obesity effect, Frankia, flavonoid, L- quebrachitol, vitamin C

The Role of Adenylyl Sulfate Reductase to Abiotic Stress in Tomato

The full-length cDNA of LeAPR1 encoded a protein of 461 amino acid residues, which contained homology with phosphoadenosine phosphosulphate reductase (PAPS reductase) in N-terminal and an adenylylsulfate reductase in N-term and C-terminal. Analysis of the deduced amino acid sequence of LeAPR1 revealed that it shares high sequence identity with potato StAPR (96% identity)(Gene bank accession no. CDC44841). We found that multiple copies of LeAPR1 gene are present in the tomato genome through southern blot using genomic DNA was digested with 3 different restriction enzymes. The expression of LeAPR1 was also examined in various organs and its expression was also detected at high levels in roots and stems. Only high amounts of LeAPR1 transcripts were detected at high transcripts in the leaves at time 0, and then reduced as the plant stressed by the NaCl and abscisic acid (ABA). After 24h treatment of NaCl and ABA were showed increasing patterns of LeAPR1 gene. Time course of LeAPR1 gene expression was examined under oxidative stresses from metyl viologen (MV) and hydrogen peroxide (). In the presence of 10 mM and MV, the levels of LeAPR1 transcript in leaves decreased after 1 h, and then increased strongly, peaked at 24 h. Our results indicated that LeAPR1 may play a role function of circadian regulation involved in abiotic stresses signaling pathways.

Lycopersicon Eculentum C 2 H 2 -type Zinc Finger Protein Induced by Oxidative Stress Especially

A tomato zinc-finger protein gene, LeZFP1, encoding the Cys2/His2-type zinc-finger transcription factor was searched from cDNA microarray analysis of gene expression following induction of the overexpressed tomato transgenic plants showing resistance for pathogen and abiotic stresses. The full-length cDNA of LeZFP1 encoded a protein of 261 amino acid residues. Analysis of the deduced amino acid sequence of LeZFP1 revealed that it shares high sequence identity with pepper CAZFP1 (81% identity). We found that single copy of LeZFP1 gene is present in the tomato genome through southern blot analysis. The LeZFP1 transcripts were constitutively expressed in the tomato mature and young leaves, but were detectable weakly in the flower, stem and root. The LeZFP1 transcripts were significantly reduced in treated leaf tissues with NaCl and mannitol. The LeZFP1 gene was induced by oxidative stress especially. Our results indicated that LeZFP1 may play a role function involved in oxidative stress signaling pathways.