• Journal of Ginseng Research
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• v.27 no.1
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• pp.17-23
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• 2003

Agrobacterium-mediated transformation of American ginseng (Panax quinquefolius L.) with strain LBA 4404 containing a rice thaumatin-like protein gene is described. The selectable markers used were phosphinothricin acetyltransferase and hygromycin phosphotransferase genes. Epicotyl explants from seedlings were precultured for 5-7 days on Murashige and Skoog medium with ${\alpha}$-naphthaleneacetic acid and 2,4 dichlorophenoxyacetic acid at 10 ${\mu}$M and 9 ${\mu}$M, respectively (ND medium), prior to Agrobacterium infection. The explants were immersed in a bacterial suspension for 20 min. A post-infection co-culture period of 3-4 days was provided on ND medium. Selection for transformed calli was conducted on ND medium with 20 mg/L phosphinothricin followed by 100 mg/L hygromycin over an 8-month period. it transformation frequency of 24.8% was achieved at the callusing phase. The presence of the transgenes in calli was confirmed by Southern hybridization and polymerase chain reaction analysis. The expression of the thaumatin-like protein gene in ginseng calli was demonstrated by Western blot analysis. Somatic embryos were produced from both transgenic calli and suspension cultures, and plantlets were recovered that expressed the transgenic thaumatin-like protein gene.

• Korean Journal of Plant Tissue Culture
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• v.28 no.5
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• pp.255-261
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• 2001

Vitis vinifera thaumatin-like protein (VVTL1) is a fruit-specific and ripening-related protein in grape. In order to isolate VVTL1-homolog gene and fruit-specific promoter from Campbell cultivar, we isolated a genomic clone containing VVTL1-homolog gene from grape genomic library through plaque hybridization. VVTL1-homolog gene has an intronless genomic structure, which the pattern is matched with those of other PR5 genes such as osmotin and osmotin-like protein genes. Transcription start site was determined by primer extension analysis. The promoter region of VVTL1-homolog gene contains a sequence or structure, especially the location and number of TCA box and ABRE (abscisic acid-responsive element), distinct from other reported plant PR5 genes, though with several known functional elements such as a TATA box and CAAT box. These results suggested that VVTL1-homolog gene may be regulated by a plant hormone, abscisic acid, and one or several stresses such osmotic pressure and pathogen infection. The isolation of fruit-specific promoter may be helpful to breed a genetically modified grape with valuable phenotype or materials in fruits.

• Food Science and Biotechnology
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• v.18 no.2
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• pp.317-322
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• 2009

The proteins from 15 types of cultivar of hot peppers cultivated in Korea were extracted and its allergenicity was investigated by immunoblotting and enzyme-linked immunosorbent assay (ELISA). The immunoblotting of hot pepper proteins extracts (HPEs) against serum of hot pepper sensitized patients revealed dominant IgE binding to 14, 37, and 40 kDa molecules. The specific levels of IgE to HPEs sample No. 1, 3, and 7 were much higher than the other samples in patients. Also, IgE binding capacity of HPEs were not reduced by thermal processing and digestion in ELISA using human IgE antibody acquired from hot pepper sensitized patients. By means of Western blotting using anti-thaumatin IgY, thaumatin-like protein (TLP) acting as allergen in several plants and fruits was detected in tested hot peppers. This study demonstrates that the antigenic protein in hot peppers are present but are differently contained according to cultivars.

• Microbiology and Biotechnology Letters
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• v.26 no.5
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• pp.379-386
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• 1998

Deterioration of food is in general caused by the presence of microorganisms and chemical compounds of food itself. There exists antimicrobial compound in the food, however, addition of food antiseptics, additives, or physico-chemical processing is a common practice. The safety of artificial chemical antiseptics became a serious public concern, therefore, new natural antiseptic compounds are in need to be developed. We have isolated a new natural antifungal protein (KBS-B2) from Astragal seed through ammonium sulfate precipitation and column chromatography using FPLC Mono-S and Superose 12HR. The purified protein inhibited growth of Candida albicans, and spore germination of food spoiling fungi such as Aspergillus ochraceus, Penicillium expensum, P. digitatum and Botrytis cineria. Antifungal effect of the KBS-B2 protein could be directly assayed by bioautography overlaying the fungal spores on the electrophoresed acrylamide gel. The comparison of N-terminal amino acid sequences of the KBS-B2 with known antifungal protein revealed that had 50% homology to thaumatin and zeamatin like proteins.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.48 no.5
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• pp.407-412
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• 2003

In order to measure an antifreezing tolerance, antifreeze proteins accumulated upon cold acclimation in apoplast were analyzed. As Dongborilho were cold-acclimated for 3 to 74 days there was an abrupt increase in apoplastic proteins up to 30 days and then decrease to the similar levels. Among the known antifreeze proteins, CLP produced in E. coli. and TLP purified from apoplast were used to generate antisera that allow to measure and localize the proteins in leaves of barley. The CLP of 27.7 kDa and TLPs of 6, 26, 27, 30, and 31 kDa were increased in their amounts in apoplast as cold treatment being longer. There was a correlation among the amounts of those proteins accumulated in apoplast and freezing tolerance as shown in field and ion leakage tests for five cultivars. The deposit of CLP was localized in the marginal area and the area adjacent to leaf vescular bundle cells in an increasing manner according to duration of cold acclimation but no variation was observed in terms of it's distribution. Based on the close correlation between levels of antifreeze proteins and degrees of freezing tolerance, the immunological methods was to develop to estimate a freezing tolerance in barley.

• Horticultural Science & Technology
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• v.31 no.3
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• pp.275-281
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• 2013

Low temperature is one of the major environmental factors that affect productivity including reduced growth and budding of vines, and changes of metabolic processes in grape (Vitis spp.). To screen the specific expression of abiotic stress-related genes against cold treatment in 'Kyoho' and 'Campbell Early' grapevines, expression of various defense-related genes was investigated by RT-PCR and real-time PCR. Among the 67 genes analyzed by RT-PCR and real-time PCR, 17 and 16 types of cDNA were up-regulated, while 5 and 6 types were down-regulated in cold-treated 'Kyoho' and 'Campbell Early' grapevines, respectively. Genes encoding carotene (Cart3564 and Cart4472), chalcone isomerase (CHI), cytochrome P450 (CYP), flavonol synthase (FLS), endo-${\beta}$-glucanase precursor (Glu), glutathione peroxidase (GPX), glutathione-S-transferase (GST), leucine-rich repeats (LRR), manganese superoxide dismutase (Mn-SOD), phenylalanine ammonia lyase (PAL), polygalacturonase-inhibiting protein (PGIP), proline rich protein 2 (PRP2), small heat shock protein (sHSP), temperature induced lipocalin (TIL), and thaumatin-like protein (TLP) were up-regulated, while those encoding CBF like transcription factor (CBF1), chitinase-like protein (CLP), cold induced protein (CIP), glycerol-3-phosphate acyltransferase (GPAT), and mitogen-activated protein kinase (MAPK) were down-regulated by low temperature treatment in both in 'Kyoho' and 'Campbell Early'.

• ALGAE
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• v.30 no.2
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• pp.147-161
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• 2015

Brown algal extracts have long been used as feed supplements to promote health of farm animals. Here, we show new molecular insights in to the mechanism of action of a fucose containing polymer (FCP) rich fraction from the brown seaweed Ascophyllum nodosum using the Caenorhabditis elegans-Pseudomonas aeruginosa PA14 infection model. FCP enhanced survival of C. elegans against pathogen stress, correlated with up-regulation of key immune response genes such as: lipases, lysozyme (lys-1), saponin-like protein (spp-1), thaumatin-like protein (tlp-1), matridin SK domain protein (msk-1), antibacterial protein (abf-1), and lectin family protein (lfp). Further, FCP caused down regulation of P. aeruginosa quorum sensing genes: (lasI, lasR, rhlI, and rhlR), secreted virulence factors (lipase, proteases, and elastases) and toxic metabolites (pyocyanin, hydrogen cyanide, and siderophore). Biofilm formation and motility of pathogenic bacteria were also greatly attenuated when the culture media were treated with FCP. Interestingly, FCP failed to mitigate the pathogen stress in skn-1, daf-2, and pmk-1 mutants of C. elegans. This indicated that, FCP treatment acted on the regulation of fundamental innate immune pathways, which are conserved across the majority of organisms including humans. This study suggests the possible use of FCP, a seaweed component, as a functional food source for healthy living.

• The Plant Pathology Journal
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• v.24 no.4
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• pp.384-391
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• 2008

Rice blast disease, caused by the pathogenic fungus Magnaporthe grisea, is a serious issue in rice (Oryza sativa L.) growing regions of the world. Transcript profiling in rice inoculated with the fungus has been investigated using the transcriptomics technology, serial analysis of gene expression (SAGE). Short sequence tags containing sufficient information which are ten base-pairs representing the unique transcripts were identified by SAGE technology. We identified a total of 910 tag sequences via the GenBank database, and the resulting genes were shown to be up-regulated in all functional categories under the fungal biotic stress. Compared to the compatible interaction, the stress and defense genes in the incompatible interaction appear to be more up-regulated. Particularly, thaumatin-like gene (TLP) was investigated in determining the gene and protein expression level utilizing Northern and Western blotting analyses, resulting in an increase in both the gene and the protein expression level which arose earlier in the incompatible interaction than in the compatible interaction.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.43 no.3
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• pp.172-178
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• 1998

Freezing-resistant plants can survive subzero temperatures by withstanding extracellular ice formation. During cold acclimation, their leaves accumulate antifreeze proteins (AFPs) that are secreted into the apoplast and have the ability to modify the normal growth of ice crystals. Three barley, two wheat and two rye cultivars were grown under two different temperature regimes (20/16$^{\circ}C$ and 5/2$^{\circ}C$, day/night). Apoplastic proteins from winter cereals were separated by SDS-PAGE and detected with antisera to AFPs from winter rye. Apoplastic proteins accumulated to much higher levels in cold-acclimated (CA) leaves compared with nonacclimated (NA) ones in winter cereals. After cold acclimation, the protein concentration of apoplastic extracts increased significantly from 0.088 $mgmL^{-1}$ to 0.448 $mgmL^{-1}$, with about 5-fold increment. Also, the apoplastic protein content per gram leaf fresh weight in CA leaves ranged from 31 $\mu\textrm{g}$ $(gFW)^{-1}$ to 120 $\mu\textrm{g}$ $(gFW)^{-1}$ with an averaged value of 77 $\mu\textrm{g}$ $(gFW)^{-1}$, and coefficients of variation of 54.9%. The CA leaves in Musketeer (a Canadian winter rye cultivar) showed the greatest AFPs and antifreeze activity followed by 'Geurumil' (a Korean winter wheat cultivar), and 'Dongbori l' (Korean facultative barley cultivar). The proteins secreted into the wheat leaf apoplast at CA condition were more numerous than those observed in winter rye, where two $\beta$-1,3-glucanase-like proteins (GLPs), two chitinase-like proteins (CLPs) and two thaumatin-like proteins (TLPs) accumulated during cold acclimation. The proteins in barley leaf apoplast at CA conditions were a little different from those in wheat leaves. The AFPs were various among and within species. More freezing-resistant cultivars had more clear and numerous bands than less freezing-resistant ones. The high determination coefficient ($R^2$ =91 %) between freezing resistance and AFPs per gram leaf fresh weight indicated that the amount of AFPs was highly related to freezing resistance in winter cereal crops.