• Journal of Ginseng Research
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• v.43 no.3
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• pp.482-487
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• 2019

Background: The mixed-cultivation of different Panax ginseng cultivars can cause adverse effects on stability of yield and quality. K-1 is a superior cultivar with good root shape and stronger disease resistance. DNA markers mined from functional genes are clearly desirable for K-1, as they may associate with major traits and can be used for marker-assisted selection to maintain the high quality of Korean ginseng. Methods: Five genes encoding pathogenesis-related (PR) proteins of P. ginseng were amplified and compared for polymorphism mining. Primary, secondary, and tertiary structures of PR5 protein were analyzed by ExPASy-ProtParam, PSSpred, and I-TASSER methods, respectively. A coding single nucleotide polymorphism (SNP)-based specific primer was designed for K-1 by introducing a destabilizing mismatch within the 3' end. Allele-specific polymerase chain reaction (PCR) and real-time allele-specific PCR assays were conducted for molecular discrimination of K-1 from other cultivars and landraces. Results: A coding SNP leading to the modification of amino acid residue from aspartic acid to asparagine was exploited in PR5 gene of K-1 cultivar. Bioinformatics analysis showed that the modification of amino acid residue changed the secondary and tertiary structures of the PR5 protein. Primer KSR was designed for specific discrimination of K-1 from other ginseng cultivars and landraces. The developed real-time allele-specific PCR assay enabled easier automation and accurate genotyping of K-1 from a large number of ginseng samples. Conclusion: The SNP marker and the developed real-time allele-specific PCR assay will be useful not only for marker-assisted selection of K-1 cultivar but also for quality control in breeding and seed programs of P. ginseng.

• The Plant Pathology Journal
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• v.29 no.1
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• pp.110-115
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• 2013

Theobroxide, a novel compound isolated from a fungus Lasiodiplodia theobromae, stimulates potato tuber formation and induces flowering of morning glory by initiating the jasmonic acid synthesis pathway. To elucidate the effect of theobroxide on pathogen resistance in plants, Nicotiana benthamiana plants treated with theobroxide were immediately infiltrated with Pseudomonas syringae pv. tabaci. Exogenous application of theobroxide inhibited development of lesion symptoms, and growth of the bacterial cells was significantly retarded. Semiquantitative RT-PCRs using the primers of 18 defense-related genes were performed to investigate the molecular mechanisms of resistance. Among the genes, the theobroxide treatment increased the expression of patho-genesis-related protein 1a (PR1a), pathogenesis-related protein 1b (PR1b), glutathione S-transferase (GST), allen oxide cyclase (AOC), and lipoxyganase (LOX). All these data strongly indicate that theobroxide treatment inhibits disease development by faster induction of defense responses, which can be possible by the induction of defense-related genes including PR1a, PR1b, and GST triggered by the elevated jasmonic acid.

• The Plant Pathology Journal
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• v.30 no.2
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• pp.208-214
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• 2014

Wild rice, Oryza grandiglumis shows hyper-resistance response to pathogen infection. In order to identify genes necessary for defense response in plants, we have carried out a subtractive hybridization coupled with a cDNA macroarray. An acidic PATHOGENESIS-RELATED1 (PR1) gene of the wild rice is highly identical to the acidic PR1 genes of different plant species. The OgPR1a cDNA has an apparent single open reading frame with a predicted molecular mass 40,621 Da and an isoelectic point of 5.14. Both in silico analysis and a transient expression assay in onion epidermal cells revealed that the OgPR1a protein could be localized in intercellular space in plants. The OgPR1a mRNA was strongly transcribed by the exogenous treatment with ethylene and jasmonic acid as well as protein phosphatase inhibitors. Additionally, ectopic expression of the OgPR1a conferred disease resistance on Arabidopsis to the bacterial and fungal infections.

• Animal cells and systems
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• v.7 no.3
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• pp.213-219
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• 2003

The response of plants to pathogens and wounding is dependent upon very sensitive perception mechanisms. Although genetic approaches have revealed a variety of resistance genes that activate common defense responses, defense-related proteins are not well characterized in plants. Therefore, we used a proteomic approach to determine which defense-related proteins are induced by salicylic acid (SA) and wounding in Chinese cabbage. We found that SA and wounding induce pathogenesis-related protein 1a (PR1a) at both protein and mRNA levels using proteomics and Northern blot analysis, respectively. This indicates that our proteomic approach is useful for identifying defense-related proteins. We also identified several other proteins that are induced by SA or wounding. Among the seven SA-induced proteins identified, four may be defense-related, including defense-related protein, phospholipase D (PLD), resistance protein RPS2 homolog, and L-ascorbate peroxidase. Out of the six wounding-induced proteins identified, three may be defense-related: heat shock cognate protein 70 (HSC70), polygalacturonase, and peroxidase P7. The precise functions of these proteins in plant defense responses await further study. However, identification of the defense-related proteins described in this study should allow us to better understand the mechanisms and signal transduction pathways involved in defense responses in Chinese cabbage.

• The Plant Pathology Journal
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• v.21 no.3
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• pp.288-292
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• 2005

Pathogenesis-related protein-1a (PR-1a) is strongly induced in tobacco plants by pathogen attack, exogenous salicylic acid (SA) application and by other developmental processes. In order to develop a rapid screening system for the selection of plant defense-inducing compounds originated from various sources, we have transformed tobacco Samsun NN plants with a chimeric construct consisting of GUS $(\beta-glucuronidase)$. In the $T_1$ generation, three transgenic lines having stable GUS expression were selected for further promoter analysis. Using GUS histochemical assay, we observed strong GUS induction driven by PR-1a promoter in PR1a-GUS transgenic tobacco leaves in response to the exogenous application of SA or benzol (1,2,3) thiadiazole-7-carbothioic acid S-methyl ester (BTH), a SA­derivative compound. In addition, GUS expression was maintained locally or systemically in PR1a-GUS transgenic line $\#5\;T_2$ generation) until after 3 days when they were treated with same chemicals. Our results suggested that the PR1a-GUS reporter gene system in tobacco plants may be applicable for the large-scale screening of defense-inducing substances.

• Korean Journal Plant Pathology
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• v.11 no.1
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• pp.53-60
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• 1995

Inoculation with the compatible strain Ds 1 of Xanthomonas campestris pv. vesicatoria caused brownish ad water-soaked lesions, but incompatible strain Bv5-4a produced hypersensitive symptoms with local necrosis on tomato (cv. Kwangyang) leaves. Bacterial populations of the compatible strains Ds 1 propagated more greatly than the incompatible strain Bv5-4a at the frist onset, but no differences were observed 5 days after inoculation. The bacterial infection induced the synthesis and accumulation of soluble proteins in tomato leaves, especially in the incompatible interaction. Native-polyacrylamide gel electrophoresis distinguished the soluble proteins in the tomato leaves infected by the compatible or incompatible strains. A protein of low molecular weight occurred only in the incompatible interaction. Some pathogenesis-related (PR) proteins, especially the 15, 18, 23, 26 and 54 kDa proteins, were detected only in the infected tomato leaves. In the two-dimensional electrophoresis, some proteins with different molecular weights (Mr. 21∼29 kDa) and the pI 8∼9 appeared more distinctly only in the incompatible interaction. These data suggest that the de novo synthesis of some PR proteins in tomato may be significant in defense against X. c. pv. vesicatoria.

• The Plant Pathology Journal
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• v.16 no.5
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• pp.264-268
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• 2000

A fluorescent material was accumulated in inoculated leaves showing necrotic local lesions of tobacco plants with N gene, Nicotiana tabacum cvs. Xanthi-nc NN, Samsun NN, Burley 21 and KF 114, and N. glutinosa, and Datura stramonium at the early growth stages by the inoculation of Tobacco mosaic virus (TMV). It was identified as a coumarin phytoalexin, scopoletin. Although the material was most prominently produced in TMV-inoculated tobacco leaves with local necrotic lesions, its accumulation was also noted in uninoculated leaves of TMV-inoculated plants. Its accumulation was somewhat greater in high resistance-induced leaves than low resistance-induced and intact leaves. Scopoletin treatment induced the expression of a pathogenesis-related protein, PR-1, prominently at the concentration of 500 or 1000 ${\mu}$g/ml. This suggests that scopoletin is a phytoalexin abundantly accumulating in N gene-containing resistant plants in response to TMV infection, and may be related to hypersensitive responses (HR) and systemic acquired resistance (SAR) in the resistant tobacco plants.

• The Plant Pathology Journal
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• v.31 no.2
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• pp.108-114
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• 2015

Curvularia lunata is an important maize foliar fungal pathogen that distributes widely in maize growing area in China, and several key pathogenic factors have been isolated. An yeast two-hybrid (Y2H) library is a very useful platform to further unravel novel pathogenic factors in C. lunata. To construct a high-quality full length-expression cDNA library from the C. lunata for application to pathogenesis-related protein-protein interaction screening, total RNA was extracted. The SMART (Switching Mechanism At 5' end of the RNA Transcript) technique was used for cDNA synthesis. Double-stranded cDNA was ligated into the pGADT7-Rec vector with Herring Testes Carrier DNA using homologous recombination method. The ligation mixture was transformed into competent yeast AH109 cells to construct the primary cDNA library. Eventually, a high qualitative library was successfully established according to an evaluation on quality. The transformation efficiency was about $6.39{\times}10^5$ transformants/$3{\mu}g$ pGADT7-Rec. The titer of the primary cDNA library was $2.5{\times}10^8cfu/mL$. The numbers for the cDNA library was $2.46{\times}10^5$. Randomly picked clones show that the recombination rate was 88.24%. Gel electrophoresis results indicated that the fragments ranged from 0.4 kb to 3.0 kb. Melanin synthesis protein Brn1 (1,3,8-hydroxynaphthalene reductase) was used as a "bait" to test the sufficiency of the Y2H library. As a result, a cDNA clone encoding VelB protein that was known to be involved in the regulation of diverse cellular processes, including control of secondary metabolism containing melanin and toxin production in many filamentous fungi was identified. Further study on the exact role of the VelB gene is underway.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.72.2-73
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• 2003

We found a soybean (Glycine max) cultivar 561 that was strongly resistant to a virulent bacterial strain of a Pseudomonas spp. Further identification revealed that the Pseudomonas spp. was a strain of Pseudomonas aeruginosa. Furthermore we identified specific genes involved in the resistance of soybean 561 and analyzed the pattern of gene expression against the Pseudomonas infection using differential-display reverse transcription PCR (DDRT-PCR). More than 126 cDNA fragments representing mRNAs were induced within 48 hours of bacteria inoculation. Among them, 28 cDNA fragments were cloned and sequenced. Twelve differentially displayed clones with open reading frames had unknown functions. Sixteen selected cDNA clones were homologous to known genes in the other organisms. Some of the identified cDNAs were pathogenesis-related genes (PR genes) and PR-like genes. These cDNAs included a putative calmodulin-binding protein, an endo-1,3-1,4-b-D-glucanase, a b-1,3-endoglucanase, a b-1,3-exoglucanase, a phytochelatin synthetase-like gene, a thiol pretense, a cycloartenol synthase, and a putative receptor-like sorineithreonine protein kinase. Among them, we found that four genes were putative pathogenesis-related genes (PR) induced significantly by the p. aeruginosa infection. These included a calmodulin-binding protein gene, a b-1,3-endoglucanase gene, a receptor-like sorine/threonine protein kinase gene, and pS321 (unknown function). These results suggest that the differentially expressed genes may mediate the strong resistance of soybean 561 to Pseudomonas aeruoginosa.

• The Plant Pathology Journal
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• v.19 no.5
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• pp.239-247
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• 2003

In Korea, a local soybean (Glycine max) genotype 56l. was found to be strongly resistant to a virulent bacterial strain of a Pseudomonas sp. SN239. Specific genes involved in the resistance of the soybean genotype 561 were identified and the pattern of gene expression against the Pseudomonas infection was analyzed using differential-display reverse transcription PCR (DDRT-PCR). More than 126 cDNA fragments representing mRNAs were induced within 48 hours of bacteria inoculation. Among them, 28 cDNA fragments were cloned and sequenced. Twelve differentially displayed clones with open reading frames had unknown functions. Sixteen selected cDNA clones were homologous to known genes of other organisms. Some of the identified cDNAs were pathogenesis-related (PR) genes and PR-like genes. These cDNAs included a putative calmodulin-binding protein; an endo-l,3-1,4-$\bate$-D-glucanase; a $\bate$-1,3-endoglucanase; a $\bate$-1,3-exoglucanase; a phytochelatin synthetase-like gene; a thiol protease; a cycloartenol synthase; and a putative receptor-like serine/threonine protein kinase. Among them, four genes were found to be putative PR genes induced significantly by the Pseudomonas infection. These included a calmodulin-binding protein gene, a $\bate$-1,3-endoglucanase gene, a receptor-like serine/threonine protein kinase gene, and pS321 (unknown function). These results suggest that the differentially expressed genes may mediate the strong resistance of soybean 561 to the strain SN239 of Pseudomonas sp.