• Research in Plant Disease
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• v.29 no.3
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• pp.286-294
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• 2023

Biological and molecular characterization of a Korean isolate of Orthotospovirus chrysanthinecrocaulis (formerly known as chrysanthemum stem necrosis virus, CSNV) isolated from Chrysanthemum morifolium was determined using host range and sequence analysis in this study. Twenty-three species of indicator plants inoculated mechanically CSNV-Kr was investigated for determination of host range. CSNV-Kr induced various local and systemic symptoms in the inoculated plant species. CSNV-Kr could not infect three plant species and induced symptomless in systemic leaves in Nicotiana tabacum cultivars, though the plant samples reacted positively with the antiserum to CSNV by double-antibody sandwich-enzyme-linked immunosorbent assay. The complete genome sequence of CSNV-Kr was determined. The L RNA of CSNV-Kr consists of 8,959 nucleotides (nt) and encodes a putative RNA-dependent RNA polymerase. The M RNA of CSNV-Kr consists of 4,835 nt and encodes the movement protein (NSm) and the glycoprotein precursor (Gn/Gc protein). The S RNA of CNSV-Kr consists of 2,836 nt and encodes NSs protein and N protein. The Gn/Gc and N sequence of CSNV-Kr were compared with those of previously published CSNV isolates originating from different countries at nucleotide and amino acid levels. The Gn/GC sequence of CSNV-Kr shared 98.8-99.5% identity with CSNV isolated from other countries and the N sequence of CSNV-Kr shared 98.8-99.6% identity. No particular region of variability could be found in either grouping of viruses. All of the CSNV isolates did not show any relationship according to geographical origins and isolation hosts, suggesting no distinct segregation of the CSNV isolates.

• The Plant Pathology Journal
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• v.22 no.4
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• pp.386-390
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• 2006

Extensive research of the Potato virus X(PVX) has been performed in in vitro transcription system using the bacteriophage T7 promoter. We constructed an efficient T-DNA based binary vector, pSNU1, and modified vectors carrying PVX replicons. The suitability of the construct to transiently express PVX RNA using Agrobacterium tumefaciens was tested by analysis of infectivity in plants. The expressed PVX RNA was infectous and systemically spread in three plant species including Nicotiana benthamiana, N. tabacum cv. Xanthi-nc, and Capsicum annuum cv. Chilsungcho. The PVX full length construct, pSPVXp31, was caused severe mosaic symptoms on N. benthamiana, severe necrotic lesions on C. annuum while milder symptoms and delayed mosaic symptoms were appeared on the systemic leaves on N. tabaccum. RT-PCR analysis confirmed the presence of PVX RNAs on both inoculated and systemic leaves in all three plant species tested. Our results indicated that PVX replicons were efficiently expressed PVX RNA in at least three tested species. Further investigation win be needed to elucidate the mechanism of PVX replication, translation, movement and assembly/disassembly processes.

• Journal of Plant Biology
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• v.38 no.3
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• pp.267-274
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• 1995

Based upon the nucleotide sequence of As strain of cucumber mosaic virus (CMV-As0 RNA4, coat protein (CP) gene was selected for the design of oligonucleotide primers of polymerase chain reaction (PCR) for detection and identification of the virus. Reverse transcription and polymerase chain reaction (RT-PCR) was performed with a set of 18-mer CMV CP-specific primers to amplify a 671 bp fragment from crude nucleic acid extracts of virus-infected leaf tissues as well as purified viral RNAs. The minimum concentrations of template viral RNA and crude nucleic acids from infected tobacco tissue required to detect the virus were 1.0 fg and 1:65,536 (w/v), respectively. No PCR product was obtained when potato virus Y-VN RNA or extracts of healthy plants were used as templates in RT-PCR using the same primers. The RT-PCR detected CMV-Y strain as well as CMV-As strain. Restriction analysis of the two individual PCR amplified DNA fragments from CMV-As and CMV-Y strains showed distinct polymorphic patterns. PCR product from CMV-As has a single recognition site for EcoRI and EcoRV, respectively, and the product from CMV-Y has no site for EcoRI or EcoRV but only one site for HindIII. The RT-PCR was able to detect the virus in the tissues of infected pepper, tomato and Chinese cabbage plants.

• Molecules and Cells
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• v.44 no.12
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• pp.911-919
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• 2021

The virus-induced genome editing (VIGE) system aims to induce targeted mutations in seeds without requiring any tissue culture. Here, we show that tobacco rattle virus (TRV) harboring guide RNA (gRNA) edits germ cells in a wild tobacco, Nicotiana attenuata, that expresses Streptococcus pyogenes Cas9 (SpCas9). We first generated N. attenuata transgenic plants expressing SpCas9 under the control of 35S promoter and infected rosette leaves with TRV carrying gRNA. Gene-edited seeds were not found in the progeny of the infected N. attenuata. Next, the N. attenuata ribosomal protein S5 A (RPS5A) promoter fused to SpCas9 was employed to induce the heritable gene editing with TRV. The RPS5A promoter-driven SpCas9 successfully produced monoallelic mutations at three target genes in N. attenuata seeds with TRV-delivered guide RNA. These monoallelic mutations were found in 2%-6% seeds among M₁ progenies. This editing method provides an alternative way to increase the heritable editing efficacy of VIGE.

• Research in Plant Disease
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• v.14 no.3
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• pp.229-232
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• 2008

An isolate of Cucumber mosaic virus (CMV), designated as Cas-CMV, was isolated from Chinese aster (Callistephus chinensis) showing severe mosaic symptom, and its properties was compared to the well-characterized Fny-CMV (subgroup IA) and As-CMV (subgroup IB) by host reaction in several indicator plants, dsRNA analysis, RT-PCR analysis, and restriction enzyme profile of the PCR products. Cas-CMV differed markedly in their host reaction to Fny-CMV or As-CMV in Cucurbita pepo cv. Black beauty. In the zucchini squash, all strains induced chlorotic spot on inoculated leaves and mosaic symptoms on upper leaves. However, symptoms induced by Cas-CMV were developed lethal necrosis on the young plants 15 to 20 days after inoculation. In experiments of dsRNA analysis and RT-PCR analysis, properties of Cas-CMV was come within subgroup I CMV. Moreover, restriction enzyme analysis using HindIII of the RT-PCR products showed that Cas-CMV belong to a member of CMV subgroup IA.

• Korean Journal Plant Pathology
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• v.10 no.3
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• pp.228-234
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• 1994

Genomic RNA was extracted from Cy strain of odontoglossum ringspot tobamovirus (ORSV-Cy) isolated from infected leaves of tobacco cv. Samsun. Size of the genomic RNA was about 6.6 kb in length. The genomic RNA was fractionated using Sephadex G-50 column chromatography into 2 fractions. They were polyadenylated at their 3'-end using E. coli poly(A) polymerase. Polyadenylated viral RNA was recovered by oligo (dT) primer adapter containing NotI restriction site and Moloney murine leukemia virus SuperScript reverse transcriptase (RNase H-). Second-strand cDNA was synthesized by using E. coli DNA ligase, E. coli DNA polymerase I and E. coli RNase H. Recombinant plasmids containing cDNAs for ORSV-Cy RNA ranged from about 800 bp to 3,000 bp. Among the selected 238 recombinants, pORCY-124 clone was the largest one covering 3'-terminal half of the viral RNA. This clone contained two restriction sites for EcoRI and XbaI and one site for AccI, AvaI, BglII, BstXI, HindIII, PstI, and TthIII 1. respectively. The clone contained partial viral replicase, a full-length movement protein and a complete coat protein genes followed by a 3' untranslated region of 414 nucleotides based on restriction mapping and nucleotide sequencing analyses. Clones pORCY-028, -068, -072, -187 and -224 were overlapped with the pORCY-124. Clones pORCY-014 and -095 covered 5' half upstream from the middle region of the viral RNA, which was estimated based on restriction mapping and partial sequence analysis. Constructed cDNA library covered more than 90% of the viral genome.

• The Plant Pathology Journal
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• v.20 no.4
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• pp.313-315
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• 2004

The coat protein gene of Korean isolate of Ricer stripe virus (RSV-Kr) was cloned and its nucleotide sequence was determined. Total RNA was extracted from infected leaves and RSV viral RNA was detected by using RT-PCR with specific primer of coat protein gene. The result of RT-PCR showed a specific band. Purified RT-PCR products of coat protein gene were ligated into the pGEM-T Easy plasmid vector and cloned cDNA was obtained for nucleotide sequence determination. Coat protein gene of RSV-Kr consisted of 969 bp long encoding a protein of 322 amino acids. RSV-Kr showed 94%-99% sequence identities to that of Japanese- and Chinese isolates.

• The Plant Pathology Journal
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• v.20 no.2
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• pp.155-157
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• 2004

We isolated Rice dwarf virus (RDV) from infected plants in rice fields (Korea, Japan, China, the Philippines and Nepal) and analyzed their genomic dsRNAs by polyacrylamide gel eletrophoresis. The genomic dsRNAs of the isolates showed distinct electrophoretic mobility profiles. The S12 coding to nonstructural protein of Korean isolate (RDV-Kr) was further analyzed by sequencing. The S12 of RDV-Kr was 1,066bp long and coded for a protein composed of 312 amino acids including three open reading frames of P12, P120Pa and P120Pb. The sequence identities were 96% and 98.6% with Japanese isolates (H, AN), 94.7% with Nepalese isolate (NEL), 94% with Chinese isolate (CK) and the Philippines isolate (P).

• The Plant Pathology Journal
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• v.27 no.4
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• pp.372-377
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• 2011

A virus causing mottle and stunt symptom on Zea mays was observed around Ulleng-do, Korea and identified as Cucumber mosaic virus (CMV-ZM) based upon biological, serological, and molecular characteristics. In host range studies, the CMV-ZM isolate produced local lesions on Datura stramonium, Vigna unguiculata, Cucurbita moschata, Chenopodium amaranticolor, Ch. quinoa, whereas this isolate produced systemic mosaic on Nicotiana tabacum cv. 'Xanthi-nc', Capsicum annuum, Solanum lycopersicum, Solanum melongena, Cucurbita pepo, and Z. mays. In addition, chlorotic local rings on inoculated leaves along with severe mosaic, malformation, and fern leaf symptoms on upper systemic leaves were shown in N. glutinosa plants. Complete nucleotide sequences of each genomic RNA segment was determined and compared to those of the other CMV strains. Comparison of the nucleotide sequence of 1a open reading frame (ORF) revealed approximately 89.2-92.4% sequence identity with each CMV subgroup IA and IB strain, while showing only 78% sequence identity with CMV subgroup II. Nucleotide sequence analysis of RNA2 ORFs revealed 85.3-97.6% sequence identity with subgroup I. In ORFs of RNA3, levels of nucleotide sequence identities were higher than 92-99.2% with CMV subgroup I and lower than 82% with CMV isolates of subgroup II. These results suggest that CMV-ZM isolate is more closely related to subgroup I than subgroup II and therefore, CMV-ZM isolate might be classified into as CMV subgroup I based on biological and molecular analysis.

• Korean Journal of Veterinary Service
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• v.42 no.2
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• pp.73-83
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• 2019

Foot-and-Mouth Disease (FMD) is a highly contagious trans-boundary viral disease caused by FMD virus, which causes huge economic losses. FMDV infects cloven hoofed (two-toed) mammals such as cattle, sheep, goats, pigs and various wildlife species. To control the FMDV, it is necessary to understand the life cycle and the pathogenesis of FMDV in host. Especially, the protein-protein interaction between FMDV and host will help to understand the survival cycle of viruses in host cell and establish new therapeutic strategies. However, the computational approach for protein-protein interaction between FMDV and pig hosts have not been applied to studies of the onset mechanism of FMDV. In the present work, we have performed the prediction of the pig's proteins which interact with FMDV based on RNA-Seq data, protein sequence, and structure information. After identifying the virus-host interaction, we looked for meaningful pathways and anticipated changes in the host caused by infection with FMDV. A total of 78 proteins of pig were predicted as interacting with FMDV. The 156 interactions include 94 interactions predicted by sequence-based method and the 62 interactions predicted by structure-based method using domain information. The protein interaction network contained integrin as well as STYK1, VTCN1, IDO1, CDH3, SLA-DQB1, FER, and FGFR2 which were related to the up-regulation of inflammation and the down-regulation of cell adhesion and host defense systems such as macrophage and leukocytes. These results provide clues to the knowledge and mechanism of how FMDV affects the host cell.