• The Plant Pathology Journal
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• 제37권2호
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• pp.99-114
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• 2021

Tan spot of wheat, caused by Pyrenophora tritici-repentis (Ptr), results in a yield loss through chlorosis and necrosis of healthy leaf tissue. The major objective of this study was to compare gene expression in resistant and susceptible wheat cultivars after infection with Ptr ToxA-producing race 2 and direct infiltration with Ptr ToxA proteins. Greenhouse experiments included exposure of the wheat cultivars to pathogen inoculum or direct infiltration of leaf tissue with Ptr-ToxA protein isolate. Samples from the experiments were subjected to RNA sequencing. Results showed that ToxA RNA sequences were first detected in samples collected eight hours after treatments indicating that upon Ptr contact with wheat tissue, Ptr started expressing ToxA. The resistant wheat cultivar, in response to Ptr inoculum, expressed genes associated with plant resistance responses that were not expressed in the susceptible cultivar; genes of interest included five chitinases, eight transporters, five pathogen-detecting receptors, and multiple classes of signaling factors. Resistant and susceptible wheat cultivars therefore differed in their response in the expression of genes that encode chitinases, transporters, wall-associated kinases, permeases, and wound-induced proteins, among others. Plants exposed to Ptr inoculum expressed transcription factors, kinases, receptors, and peroxidases, which are not expressed as highly in the control samples or samples infiltrated with ToxA. Several of the differentially expressed genes between cultivars were found in the Ptr resistance QTLs on chromosomes 1A, 2D, 3B, and 5A. Future studies should elucidate the specific roles these genes play in the wheat response to Ptr.

• Journal of Microbiology and Biotechnology
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• 제31권2호
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• pp.250-258
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• 2021

Among various species of marine bacteria, those belonging to the genus Halomonas have several promising applications and have been studied well. However, not much information has been available on their antibiotic resistance. In our efforts to learn about the antibiotic resistance of strain Halomonas socia CKY01, which showed production of various hydrolases and growth promotion by osmolytes in previous study, we found that it exhibited resistance to multiple antibiotics including kanamycin, ampicillin, oxacillin, carbenicillin, gentamicin, apramycin, tetracycline, and spectinomycin. However, the H. socia CKY01 resistance pattern to kanamycin, gentamicin, apramycin, tetracycline, and spectinomycin differed in the presence of 10% NaCl and 1% NaCl in the culture medium. To determine the mechanism underlying this NaCl concentration-dependent antibiotic resistance, we compared four aminoglycoside resistance genes under different salt conditions while also performing time-dependent reverse transcription PCR. We found that the aph2 gene encoding aminoglycoside phosphotransferase showed increased expression under the 10% rather than 1% NaCl conditions. When these genes were overexpressed in an Escherichia coli strain, pETDuet-1::aph2 showed a smaller inhibition zone in the presence of kanamycin, gentamicin, and apramycin than the respective control, suggesting aph2 was involved in aminoglycoside resistance. Our results demonstrated a more direct link between NaCl and aminoglycoside resistance exhibited by the H. socia CKY01 strain.

• 한국토양비료학회지
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• 제44권1호
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• pp.53-57
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• 2011

The MuSI is known as a multiple stress resistant gene with several lines. A previous study using RT-PCR showed that the expression of MuSI gene in tobacco plant induced its tolerance to Cd stress. This study was conducted to examine the enhanced Cd tolerance of the MuSI transgenic tobacco plant through germination test and to understand the role of the involved antioxidant enzymes for the exhibited tolerance. Germination rate of MuSI transgenic tobacco was more than 10% higher than that of wild-type tobacco, and seedlings of MuSI transgenic tobacco grew up to 1.6 times larger and greener than seedlings of wild-type tobacco at 200 and 300 ${\mu}M$ Cd. From the third to the fifth day, CAT activities at 100 and 200 ${\mu}M$ Cd and APX activities at 100, 200 and 300 ${\mu}M$ Cd of MuSI transgenic tobacco were up to two times higher than those of wild-type tobacco. MuSI gene is shown to enhance the activities of antioxidant enzymes resulting in higher tolerance to oxidative stress compared with the control plant.

• 한국축산식품학회지
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• 제26권3호
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• pp.394-401
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• 2006

Forty Clostridium perfringens isolates were obtained from twelve animal products, following the examination of eighty six beef, pork, broiler chicken and salami meat products, and eleven milk powder products. There were 21 isolates from salami stored at $25^{\circ}C$, 3 isolates from pork, 4 isolates from beef, 9 isolates from broiler chicken, and 3 isolates from milk powder. Only the cpa gene encoding a toxin among the 5 toxin genes tested (cpa, cpb, etx, iap, and cpe) was detected in all forty isolates, suggesting contamination with C. perfringens type A. DNA fingerprinting analysis using PCR of the tRNA intergenic spacer (tDNA-PCR) and the 16S-23S internal transcribed spacer (ITS-PCR), and randomly amplified polymorphic DNA (RAPD) analysis were attempted to differentiate the isolates. RAPD analysis was the most discriminating method among the three PCR analyses. Isolates from the same products tended to show similar RAPD patterns. Antimicrobial susceptibility tests showed that some isolates from broiler chickens had the same antibiogram with multiple resistance to streptomycin, colistin, and ciprofloxacin. Antibiograms were similar between isolates from the same livestock products, but differed considerably between the products.

• The Plant Pathology Journal
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• 제17권2호
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• pp.88-93
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• 2001

Oxidative stress is one of the major limiting factor in plant productivity. Reactive oxygens species (ROS) generated during metabolic processes damage cellular functions and consequently lead to disease, senescence and cell death. Plants have evolved an efficient defense system by which the ROS is scavenged by antioxidant enzymes such as superoxide dismutase (SOD) and ascorbate peroxidase (APX). Attempts to reduce oxidative damages under the stress conditions have included the manipulation of 갠 scavenging enzymes by gene transfer technology. Increased SOD activities of transgenic plants lead to increased resistance against oxidative stresses derived from methyl viologen (MV), and from photooxidative damage caused by high light and low temperature. Transgenic tobacco plants overexpressing APX showed reduced damage following either MV treatment of photooxidative treatment. Overexpression of glutathion reductase (GR) leads to increase in pool of ascorbate and GSH, known as small antioxidant molecules. These results indicate through overexpression of enzymes involved in ROS-scavenging could maintain or improve the plant productivities under environment stress condition. In this study, the rational approaches to develop stress-tolerant plants by gene manipulation of antioxidant enzymes will be introduced to provide solutions for the global food and environmental problems in the $21^\textrm{st}$ century.

• 한국동물위생학회지
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• 제32권1호
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• pp.61-76
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• 2009

At the present study, it was aimed to explore the molecular genetic characterization of multiple antimicrobial resistant Salmonella spp. isolates from pigs and cattle. A total of 138 Salmonella Typhimurium (S. Typhimurium) isolates were typed with phage, among them, 83.3% of S. Typhimurium tested could divide into a 10 phage types. Definitive type 193 (DT193) (25.4%) and DT195 (24.6%) were exhibited as the dominant types. DT104 and U302 were found from pigs and cattle. On the other hand, S. Enteritidis had 6 phage types, of them, phage type 21 (PT21) and PT11b were the popular types. In the plasmid profiles, 135 of S. Typhimurium isolates were exhibited 1 to 6 plasmid bands which molecular weight ranged from 90 to 2kb. 35 isolates (25.4%) harbored a 90kb plasmid which is thought to be the serotype specific virulence plasmid. Two of twenty five S. Enteritidis had common plasmids at 2 and 1.5kb. With multiplex polymerase chain reaction, virulence genes (invA and spvC) were detected from all Salmonella spp. from 167 of S. Typhimurium, S. Enteritidis and chloramphenicol resistant S. Schwarzengrund, but some drug resistant genes, such as PSE-1, cml/tetR and flo were not determined but other drug resistant genes, for example TEM and int were found. The detection rates of spvC, TEM and int gene was 35.3%, 29.3% and 72.5%, respectively. The TEM gene was highly popular in S. Typhimurium, which was detected from ampicillin and amoxicillin resistant strains as 95.9%. int gene was able to detect from all the isolates identified as multidrug resistsnt (MDR), particularly DT193 was thought as the most prevalent virulence and multidrug resistance isolate. The major plasmid profile and drug resistance pattern of DT193 were 90, 40, 10.5, 6.3, 3.0kb and ACCbDNaPSSuT, respectively. MDR was commonly found in other phage types, particularly DT104, U302 and DT203.

• Journal of Microbiology and Biotechnology
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• 제19권10호
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• pp.1265-1270
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• 2009

The objectives of this study were to determine the antibiotypes, SCCmec subtypes, PVL carriage, and genetic diversity of MRSA strains from a tertiary hospital. Sixty-six MRSA strains were selected randomly (2003, 2004, and 2007) and tested for the Panton-Valentine leukocidin gene, mecA gene, and SCCmec type via a PCR. The antibiograms were determined using a standard disc diffusion method, and the genetic diversity of the isolates was determined by PFGE. Thirty-four antibiograms were obtained, with 55% of the 66 strains exhibiting resistance to more than 4 antimicrobials. All the isolates remained susceptible to vancomycin, and low resistance rates were noted for fusidic acid (11%), rifampicin (11%), and clindamycin acid (19%). The MRSA isolates that were multisensitive (n=12) were SCCmec type IV, whereas the rest (multiresistant) were SCCmec type III. Only two isolates (SCCmec type IV) tested positive for PVL, whereas all the isolates were mecA-positive. The PFGE was very discriminative and subtyped the 66 isolates into 55 pulsotypes (F=0.31-1.0). The multisensitive isolates were distinctly different from the multidrug-resistant MRSA. In conclusion, no vancomycin-resistant isolate was observed. The Malaysian MDR MRSA isolates were mostly SCCmec type III and negative for PVL. These strains were genetically distinct from the SCCmec type IV strains, which were sensitive to SXT, tetracycline, and erythromycin. Only two strains were SCCmec IV and PVL-positive. The infections in the hospital concerned were probably caused by multiple subtypes of MRSA.

• 한국육종학회지
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• 제51권3호
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• pp.263-276
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• 2019

'신진백'은 농촌진흥청 국립식량과학원에서 벼흰잎마름병에 대한 저항성원을 다양화하고 저항성 유전자 집적을 통해 저항성이 향상된 고품질 중만생 벼 품종을 개발하고자 육성되었다. 벼흰잎마름병 저항성 유전자 Xa3와 xa5를 가지고 있는 '익산493호'(품종명 '진백')를 모본으로 하고 최고품질 품종이면서 Xa3를 보유하고 있는 '호품'과 야생벼 O. longistaminata에서 유래한 저항성 유전자 Xa21를 자포니카 우량계통 '수원345호' 유전배경으로 도입한 근동질 계통 'HR24670-9-2-1'간 F₁을 부본으로 삼원교배하여 육성되었다. 계통육성과정 중 벼흰잎마름병에 대한 저항성 생물검정과 분자표지를 활용한 저항성 유전자 도입여부 확인을 통해 저항성 유전자가 집적된 계통을 선발하여 생산력 검정시험과 지역적응성 검정시험을 거쳐 개발되었다. '신진백'은 보통기 보비재배에서 평균 출수기 8월 19일로 '남평'에 비해 4일 늦은 중만생종으로 간장이 70 cm로 '남평'보다 6 cm 작은 단간 내도복 품종이다. '신진백'은 우리나라 최초로 Xa21 저항성 유전자가 도입되었으며, 세 개의 저항성 유전자 Xa3, xa5, Xa21이 집적된 실용적인 재배품종으로 병원성이 강한 K3a 균계를 포함하여 우리나라 벼흰잎마름병균에 광범위 고도 저항성을 반응을 나타냈다. 또한 도열병과 줄무늬잎마름병에도 강한 복합 내병성으로 친환경 재배 적성을 갖추고 있다. '신진백'은 '남평'과 비슷한 수량성을 나타냈다. '신진백'은 쌀의 외관품위가 좋고 밥맛이 양호하며 도정 특성이 우수하여 벼흰잎마름병 저항성 품종의 품질 향상에 기여하였다. '신진백'은 새로운 벼흰잎마름병 저항성 유전자 Xa21를 열악형질 수반없이 재배품종으로 도입하였으며 세 개의 저항성 유전자를 집적함으로써 벼흰잎마름병균에 광범위 고도 저항성을 확보한 복합 내병성 품종으로 벼흰잎마름병 발병상습지 재배에 적합하며 벼흰잎마름병 저항성 향상을 위한 육종사업에 활용되고 있다(품종보호권 등록번호: 제7273호; 2018. 6. 25.).

• Journal of Plant Biotechnology
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• 제43권3호
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• pp.317-331
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• 2016

Development of disease resistant plant is one of the important objectives in rice breeding programs because biotic stresses can adversely affect rice growth and yield losses. This study was conducted to identify lines with multiple-resistance genes to biotic stress among 173 hybrid rice breeding lines and germplasms using DNA-based markers. Our results showed that one hybrid rice line [IR98161-2-1-1-k1-3 (IR86409-3-1-1-1-1-1/IRBB66)] possessed 5 bacterial blight resistance genes (Xa4, xa5, Xa7, Xa13 and Xa21) while two hybrid rice lines [IR98161-2-1-1-k1-2 (IR86409-3-1-1-1-1-1/IRBB66) and 7292s (IR75589-31-27-8-33S(S1)/IR102758B)] possessed 3 bacterial blight resistance genes (Xa4, Xa7 and Xa21, and Xa3, Xa4 and xa5). Molecular survey on rice blast disease revealed that most of these lines had two different resistant genes. Only 11 lines possessed Pib, Pi-5, and Pi-ta. In addition, we further surveyed the distribution of insect resistant genes, such as Bph1, Bph18(t), and Wbph. Three hybrid breeding lines [IR98161-2-1-1-k1-3 (IR86409-3-1-1-1-1-1/IRBB66), IR98161-2-1-1-k1-2 (IR86409-3-1-1-1-1-1/IRBB66), and 7292s (IR75589-31-27-8-33S(S1) /IR102758B)] contained all three resistance genes. Finally, we obtained four hybrid rice breeding lines and germplasms [IR98161-2-1-1-k1-2 (IR86409-3-1-1-1-1-1/IRBB66), Damm-Noeub Khmau, 7290s, and 7292s (IR75589-31-27-8-33S(S1)/IR102758B)] possessing six-gene combination. They are expected to provide higher level of multiple resistance to biotic stress. This study is important for genotyping hybrid rice with resistance to diverse diseases and pests. Results obtained in this study suggest that identification of pyramided resistance genes is very important for screening hybrid rice breeding lines and germplasms accurately for disease and pest resistance. We will expand their cultivation safely through bioassays against diseases, pests, and disaster in its main export countries.

• Journal of Microbiology and Biotechnology
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• 제27권2호
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• pp.335-341
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• 2017

The complexity of the bacterial recombination system is a barrier for the construction of bacterial mutants for the further functional investigation of specific genes. Several protocols have been developed to inactivate genes from the genus Pseudomonas. Those protocols are complicated and time-consuming and mostly do not enable easy construction of multiple knock-ins/outs. The current study describes a single and double crossover-recombination system using an optimized vector-free allele-exchange protocol for gene disruption and gene replacement in a single species of the family Pseudomonadaceae. The protocol is based on self-ligation (circularization) for the DNA cassette which has been obtained by overlapping polymerase chain reaction (Fusion-PCR), and carries an antibiotic resistance cassette flanked by homologous internal regions of the target locus. To establish the reproducibility of the approach, three different chromosomal genes (ncRNA31, rpoN, rpoS) were knocked-out from the root-associative bacterium Pseudomonas stutzeri A1501. The results showed that the P. stutzeri A1501 mutants, which are free of any plasmid backbone, could be obtained via a single or double crossover recombination. In order to optimize this protocol, three key factors that were found to have great effect on the efficiency of the homologous recombination were further investigated. Moreover, the modified protocol does not require further cloning steps, and it enables the construction of multiple gene knock-in/out mutants sequentially. This work provides a simple and rapid mutagenesis strategy for genome editing in P. stutzeri, which may also be applicable for other gram-negative bacteria.