• Journal of Microbiology and Biotechnology
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• v.23 no.1
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• pp.22-28
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• 2013

Xo2276 is a putative transcription activator-like effector (TALE) in Xanthomonas oryzae pv. oryzae (Xoo). Xo2276 was expressed with a TAP-tag at the C-terminus in Xoo cells to enable quantitative analysis of protein expression and secretion. Nearly all TAP-tagged Xo2276 existed in an insoluble form; addition of rice leaf extracts from a Xoosusceptible rice cultivar, Milyang23, significantly stimulated secretion of TAP-tagged Xo2276 into the medium. In a T3SS-defective Xoo mutant strain, secretion of TAPtagged Xo2276 was blocked. Xo2276 is a Xoo ortholog of Xanthomonas campestris pv. vesicatoria (Xcv) AvrBs3 and contains a conserved DNA-binding domain (DBD), which includes 19.5 tandem repeats of 34 amino acids. Xo2276- DBD was expressed in E. coli and purified. Direct in vitro recognition of Xo2276-DBD on a putative target DNA sequence was confirmed using an electrophoretic mobility shift assay. This is the first study measuring the homologous expression and secretion of Xo2276 in vitro using rice leaf extract and its direct in vitro binding to the specific target DNA sequence.

• 한국균학회소식:학술대회논문집
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• 2015.05a
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• pp.61-61
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• 2015

Rice blast, caused by the fungal pathogen Magnaporthe oryzae, is one of the most destructive diseases of rice worldwide. The rice - M. oryzae pathosystem has become a model in the study of plant - fungal interactions due to its economic importance and accumulating knowledge. During the evolutionary arms race with M. oryzae, rice plants evolved a repertoire of Resistance (R) genes to protect themselves from diseases in a gene-for-gene fashion. M. oryzae secretes a battery of small effector proteins to manipulate host functions for its successful infection, and some of them are recognized by host R proteins as avirulence effectors (AVR), which turns on strong immunity. Therefore, the analysis of interactions between AVRs and their cognate R proteins provide crucial insights into the molecular basis of plant - fungal interactions. Rice blast resistance genes Pik, Pia, Pii comprise pairs of protein-coding ORFs, Pik-1 and Pik-2, RGA4 and RGA5, Pii-1 and Pii-2, respectively. In all three cases, the paired genes are tightly linked and oriented to the opposite directions. In the AVR-Pik/Pik interaction, it has been unraveled that AVR-Pik binds to the N-terminal coiled-coil domain of Pik-1. RGA4 and RGA5 are necessary and sufficient to mediate Pia resistance and recognize the M. oryzae effectors AVR-Pia and AVR1-CO39. A domain at the C-terminus of RGA5 characterized by a heavy metal associated domain was identified as the AVR-binding domain of RGA5. Similarly, physical interactions among Pii-1, Pii-2 and AVR-Pii are being analyzed.

• The Plant Pathology Journal
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• v.32 no.1
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• pp.8-15
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• 2016

To identify a novel biopesticide controlling rice blast disease caused by Magnaporthe oryzae, 700 plant extracts were evaluated for their inhibitory effects on mycelial growth of M. oryzae. The L. foenum-graecum Herba extract showed the lowest inhibition concentration ($IC_{50}$) of $39.28{\mu}g/ml$, which is lower than the $IC_{50}$ of blasticidin S ($63.06{\mu}g/ml$), a conventional fungicide for rice blast disease. When treatments were combined, the $IC_{50}$ of blasticidin S was dramatically reduced to $10.67{\mu}g/ml$. Since ABC transporter genes are involved in fungicide resistance of many organisms, we performed RT-PCR to investigate the transcriptional changes of 40 ABC transporter family genes of M. oryzae treated with the plant extract, blasticidin S, and tetrandrine, a recognized ABC transporter inhibitor. Four ABC transporter genes were prominently activated by blasticidin S treatment, but were suppressed by combinational treatment of blasticidin S with the plant extract, or with tetrandrine that didn't show cellular toxicity by itself in this study. Mycelial death was detected via confocal microscopy at 24 h after plant extract treatment. Finally, subsequent rice field study revealed that the plant extract had high control efficacy of 63.3% and should be considered a biopesticide for rice blast disease. These results showed that extract of L. foenum graecum Herba suppresses M. oryzae ABC transporter genes inducing mycelial death and therefore may be a potent novel biopesticide.

• Microbiology and Biotechnology Letters
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• v.29 no.1
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• pp.12-17
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• 2001

Aspergil-lus coreanus NR-15 and Aspergilus oryzae NR-2-5 from traditional Korean Nuruk were selected as parental strains producing starch hydrolysis enzyme. Xll(Arginine-) mutant from A. coreanus NR 15-1 showed high glu-doamylase activity and total acid productivity. Z6(Adenine-) mutant from A. oryzae NR2-5 showed the highest $\alpha$-amylase activity. Therefore, both XII and Z6 mutants were selected and investigated for the optimal conditions of protoplast formation for protoplast fusion. Mixture of equal amount of cellulase and driselase(10mg/ml each) was the most effective as lytic enzymes. The optimal pH and temperature for protoplast formation were 5.0 and $30^{\circ}C$, respectively. The most effective reaction for protoplast formation time was 4 hours. The maximum of protoplst for- mation of Xll mutant and Z6 mutant were $6.54$\times$10^{7}$ protoplasts/ ml and $3.04$\times$10^{ 7}$ protoplasts/ml, and the regen-eration frequencies of the protoplasts were 11.3% and 11.6%, respectively. The size of the protoplasts from X11 and Z6 mutants were 3~6 $\mu\textrm{m}$ and 4~9$\mu\textrm{m}$, respectively.

• The Plant Pathology Journal
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• v.29 no.4
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• pp.435-439
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• 2013

Rice blast fungus, Magnaporthe oryzae, inflicts serious damage to global rice production. Due to high variability of this fungal pathogen, resistance of newly-released rice cultivars is easily broken down. To understand the population structure of M. oryzae, we analyzed the genetic diversity of the Korean population using multilocus microsatellite typing. Eleven microsatellite markers were applied to the population of 190 rice isolates which had been collected in Korea for two decades since the 1980's. Average values of gene diversity and allele frequency were 0.412 and 6.5, respectively. Comparative analysis of the digitized allele information revealed that the Korean population exhibited a similar level of allele diversity to the integrated diversity of the world populations, suggesting a particularly high diversity of the Korean population. Therefore, these microsatellite markers and the comprehensive collection of field isolates will be useful genetic resources to identify the genetic diversity of M. oryzae population.

• Microbiology and Biotechnology Letters
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• v.49 no.2
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• pp.217-224
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• 2021

Rice blast caused by Pyricularia oryzae, which is a major threat to food security worldwide, markedly decreases the yield of rice. Some rhizobacteria called 'plant growth-promoting rhizobacteria' inhibit plant pathogens and improve plant growth by secreting iron-chelating siderophores. The decreased availability of iron adversely affects the survival of pathogens, especially fungal pathogens, in the rhizosphere. This study aimed to determine the morphological diversity of siderophore-producing bacteria, analyze the type of siderophores produced by the bacteria, and examine their growth-inhibitory activity against Pyricularia oryzae. The rhizobacteria were isolated from the rhizosphere of Sembada Hitam variety of black rice plants in Pakem, Sleman, Yogyakarta, Indonesia. In total, 12 distinct isolates were screened for the production of siderophores. It was found that 9 out of 12 bacteria produced siderophore and most of them were Gram positive bacteria. The best siderophore-producing isolates with different type of siderophore were used in further studies. The IS3 and IS14 isolates were found to be the best siderophore producer that produced hydroxamate and mixed type of hydroxamate-carboxylate type of siderophore, respectively. In the dual culture assay, IS14 showed a strong antagonistic effect against Pyricularia oryzae by the 81.17% inhibition.

• Journal of Applied Biological Chemistry
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• v.54 no.4
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• pp.231-237
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• 2011

A gene encoding a putative alanine racemase in Xanthomonas. oryzae pv. oryzae was cloned, expressed and characterized. Expression of the cloned gene was performed in Escherichia coli BL21(DE3)pLys using a pET-21(a) vector harbouring $6{\times}histidine$ tag. Purification of the recombinant alanine racemase by affinity chromatography resulted in major one band by sodium dodecyl sulfate polyacryl amide gel electrophoresis analysis, showing about 45 kDa of molecular weight. The alanine racemase gene, cloned in this experiment, appears to be constitutively expressed in X. oryzae, as analyzed by reverse transcriptase polymerase chain reaction. The enzyme was the most active toward L-alanine and secondly D-alanine, showing a racemic reaction, thus the enzyme is considered as an alanine racemase. The enzyme was considerably activated by addition of pyridoxal-5-phosphate (PLP), showing that 75% increase in activity was observed at 0.3 mM, compared with control. D-Cysteine as well as L-cysteine significantly inhibited the enzyme activity. The inhibitions by cysteines were more prominent in the absence of PLP, showing 9 and 5% of control activity at 2 mM of addition, respectively. The enzyme was the most active at pH 8.0 and more stable at alkaline pHs than acidic pH condition.

• Korean Journal of Microbiology
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• v.17 no.2
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• pp.58-64
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• 1979

Studies on some properties of cellulase isolated from Piricularia oryzae. Crude cellulases were prepared from dried rice plant powder (Tong-il, Pal-dal) culture of P. oryzae(N-2, C-8, T-2). The best yield of enzyme was obtained from the medium using Tong-il rice plant powder for P. oryzae cav. N-2 and 2%-sucrose concentration in preculture media. Two units of the enzyme were incubated at $60^{\circ}C$ for 1 hour with 1.0ml, 0.6% Na-CMC. The optimum temperature for the enzyme activity was at $60^{\circ}C$ and the optimum pH was at pH4.0. When Na-CMC was used as substrate the $K_m$ values of crude enzyme were calculated to be $1.05{\times}10^{-4}\;mM\;and\;V_{max}$ was 2.8 mmole/hour. A 10-fold partial purification was achieved by $(NH_4)_2SO_4$ precipitation followed by column chromatography on DEAE Sephadex A-25.

• Korean Journal of Food Science and Technology
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• v.36 no.1
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• pp.81-85
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• 2004

Quality characteristics of kochujang made with Aspergillus oryzae-and/or Cordyceps sp.-inoculated koji were investigated. Protease activity of Cordyceps sp.-inoculated koji was higher than that of A. oryzae-inoculated one. Sensory evaluation showed that kochujang made with mixture of A. oryzae-and Cordyceps sp.-inoculated koji (70 : 30, w/w) was superior to others.

• Journal of the Korean Society of Food Science and Nutrition
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• v.13 no.3
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• pp.238-246
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• 1984

The objective of this experiment was to improve the quality of Kochujang and to shorten the aging time of it by using the Kojis of various Aspergillus strains. During the autumn season Kochujangs were mashed with Koji of Aspergillus oryzae, Aspergillus kawachii and Aspergillus shirousamii. Chemical composition of the Kochujang was determined during the aging period. There was no significant difference in quality and component between the Kochujang made with Aspergillus oryzae cultured Koji and Aspergillus kawachii cultured Koji. But the Kochujang with Aspergillus shirousamii cultured Koji was better in flavor, color and amino nitrogen contents than the Kochujang with Aspergillus oryzae cultured Koji, though two Kochuangs were almost the same in general components. The significant difference between the two tested Kochuangs was observed in pH, and the pH of the Kochujong with Aspergillus shirousamii cultured Koji was extremely low during the aging period. Thus it is presumed that Aspergillus shirousamii Koji can be used for the improvement of quality of Kochujang if a suitable method is applied to prevent pH from lowering.