• The Plant Pathology Journal
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• v.19 no.3
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• pp.171-176
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• 2003

The complete nucleotide sequences of the genomic RNAs of Soybean mosaic virus strains GS (SMV-G5) and G7H (SMV-G7H) were determined and compared with sequences of other SMV strains. Each viral RNA was determined to be 9588 nucleotides in length excluding the poly (A) tail and contained an open reading frame to encode a polyprotein subsequently processed into up to ten proteins by proteolytic cleavage. Com-parison of the amino acid sequences with those of other SMV strains showed high percentage of amino acid sequence homology with the same genome organization. The nucleotide and the deduced amino acid sequences between SMV-G5 and SMV-G7H were greater than 99% identity. When compared with those of other SMV strains in a phylogenetic analysis of the nucleotide and deduced amino acid sequences, they formed a distinct virus clade showing over 97％ amino acid identity, but were more distantly related to the other potyvirus (44.1-69.6% identity). Interestingly, SMV G7H strain caused a severe mosaic or necrosis symptom in soybean cultivars including Jinpum-1, Jinpum-2, and Sodam, whereas, no symptom was observed in SMV-G5 inoculation. Complete nucleotide sequences of these strains will give clues for determining symptom determinant(s) in future research.

• Applied Biological Chemistry
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• v.37 no.1
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• pp.56-63
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• 1994

Oligonucleotides for a conserved region of the coat protein gene of cucumber mosaic virus (CMV) and a hammerhead structure ribozyme against CMV RNA were synthesized using a DNA synthesizer. Both strands of oligonucleotides were annealed and restricted with BamHI/SacI, then cloned into a plasmid pBS SK (+). The cloned CMV substrate and ribozyme were sequenced to verify correct constructions. In vitro transcriptions were carried out by using T7 RNA polymerase with BssHII or SspI digests of $1\;{\mu}g$ of substrate and ribozyme clones. The size of substrate RNA was 176 nucleotides (nt) containing 50 nt of CMV RNA sequence, 6 nt of XbaI restriction site and 120 nt of vector-derived sequence in the case of BssHII digest. The size of ribozyme RNA was 164 nt containing 40 nt of ribozyme RNA sequence and same sequences of substrate. Substrate RNA was efficiently cleaved into two fragments (96 nt and 80 nt) by ribozyme RNA. This endonucleolytic cleavage occurred more efficiently at $55^{\circ}C$ than $37^{\circ}C$. SspI digest-derived substrate RNA (2234 nt) was also cleaved into two fragments by the same ribozyme. SspI digest-derived ribozyme RNA (2222 nt) cleaved the above substrate to two fragments. In vitro-tested ribozyme construct is being cloned into a plant transformation vector to develop virus-resistant plants.

• The Plant Pathology Journal
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• v.35 no.1
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• pp.32-40
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• 2019

Symptoms of fig mosaic disease have been noticed on leaves of fig (Ficus carica) for several decades, in Montenegro. In 2014, leaf samples were collected from trees of six fig cultivars in a plantation located in the main fig-producing area of Montenegro, to study the disease. After RNA isolation, samples were tested by RT-PCR for detection of nine fig viruses and three viroids. Four viruses were detected: fig leaf mottle-associated virus 1 (FLMaV-1), fig mosaic virus (FMV), fig mild mottle-associated-virus (FMMaV) and fig badnavirus 1 (FBV-1). Most of the viruses were present in mixed infections. The amplicons of the viruses were directly sequenced from both directions. A BLAST search of these sequences revealed sequence identities with their closest counterparts at GenBank of 92, 97, 92 and 100%, for FLMaV-1, FMV, FMMaV and FBV-1, respectively. Different responses in symptom expression due to the various virus combinations detected have been demonstrated. Variety $Su{\check{s}}ilica$ had the least symptom expression, with only one virus (FBV-1) found. Considering that the production of figs in Montenegro is increasing and has a substantial relevance in this geographic location, the results indicate that more attention should be given to improving the phytosanitary condition of fig trees in the country.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.139.2-140
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• 2003

Ornithogalum showing mosaic symptoms were collected from the isolated field of National Plant Quarantine Service in Sengrimmyon of Kyungnam province. Electron microscopic examination of negatively strained preparation was filamentous particle of 740nm in lenght. Indirect-ELISA determined that the virus was serologically related to potyvirus. A single major protein band of Mr 30,000 was observed after sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Indicator plant test showed masaic, necrotic local lesion and sunken areas in leaves of Nicotiana clevelandii and Tetragonia expansa, while the others of indicator plants did not infect. An enzyme-aided purification protocal was used, which eliminated a highly viscous mucilage from extracts of the Omithogalum. Total RNA extracted from infected Omithogalum leaves were amplified of 411b.p fragment in reverse transcription (RT)-PCR when primers specilic for the coat protein gene. An isolate of Omithogalum mosaic virus (OrMV) of the genus Potyvirus was identified as the casual agent of the disease on the basis of electron microscopic, biological and serological reaction.

• Journal of Apiculture
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• v.32 no.4
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• pp.281-293
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• 2017

Sacbrood virus (SBV) is one of the main pathogenic RNA viruses of honeybee. SBV is found worldwide and many local strains have been reported, such as kSBV, cSBV, and wSBV. In this study, SBV-specific DNA fragments were cloned and sequenced by reverse-transcription PCR from 4 populations of A. mellifera, 4 sequences from 1 population belonged to the 2134D51 genotype (349 nucleotides, nt) and 12 sequences from 3 populations belonged to the 2100D0 genotype (400 nt) among the 16 determined sequences. A total of 87 points of mismatches were found by comparison with the most similar sequences in GenBank. Seventeen single-nucleotide polymorphisms (SNP) were detected, and 6 SNP-patterns in the 2100D0 genotype and 2 SNP-patterns in the 2134D51 genotype were identified based on SNP positions. In SNP-pattern 2, 10 SNPs were detected, but only 2 SNPs were found in SNP-pattern7. Meanwhile, one SNP-pattern was found from one RNA-sample, multi SNP-patterns were detected from other RNA-samples. Large numbers of SNP variants indicate that vast numbers of point-mutations on SBV have occurred since SBV invaded Korea and that SNP smay have been introduced individually over time. Thorough analysis of SNP variants will not only define the local infection-route, but also the relationships between SNP-pattern and SBV-pathogenic abilities.

• The Plant Pathology Journal
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• v.22 no.2
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• pp.139-146
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• 2006

The 3' region of Potato virus X (PVX) has the 74 nt 3'-nontranslated region (NTR) that is conserved among all potexviruses and contains several cis-acting elements for minus-strand and plus-strand RNA accumulation. Three stem-loop structures (SL1-SL3), especially formation of SL3 and U-rich sequence of SL2, and near upstream elements in the 3' NTR were previously demonstrated as important cis-acting elements. To Investigate the binding of these cis-acting elements within 3' end with host protein, we used the electrophoretic mobility shift assays (EMSA) and UV-cross linking analysis. The EMSA with cellular extracts from tobacco and RNA transcripts corresponding to the 150 nt of the 3' end of PVX RNA showed that the 3' end of PVX formed complexes with cellular proteins. The specificity of protein binding was confirmed through competition assay by using with 50-fold excess of specific and non-specific probes. We also conducted EMSA with RNAs containing various mutants on those cis-acting elements (${\Delta}10$10, SL3B, SL2A and ${\Delta}21$; J Mol Biol 326, 701-720) required for efficient PVX RNA accumulation. These analyses supported that these cis-acting elements are required for interaction with host protein(s). UV-cross linking analysis revealed that at least three major host proteins of about 28, 32, and 42 kDa in mass bound to these cis-elements. These results indicate that cis-acting elements from 3' end which are important for minus and plus-strand RNA accumulation are also required for host protein binding.

• Korean Journal of Veterinary Service
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• v.35 no.3
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• pp.169-174
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• 2012

Japanese encephalitis virus (JEV) is one of the main causes of viral encephalitis in human and animals. For over 30 years, a live attenuated JEV vaccine strain has been used in the veterinary field, and it is required to conduct quality evaluation studies on the commercial vaccines. For the quality control of live attenuated JEV vaccine, we investigated the nucleotide sequence similarity of prME gene derived from five JEV vaccines commercially available in pigs in Korea. The Vero cells infected with JEV vaccines showed specific cytopathic effect, which was characterized by rounding and detached cells. In the phylogenetic analysis, all of the vaccine strains showed a close relationship with the original vaccine seed strain (Anyang 300) and clustered into the genotype 3. In comparison of the nucleotide and deduced amino acid sequences of prME genes with the original strain, all JEV vaccine strains showed high amino acid similarity ranging from 98.9% to 99.5%, but had several point mutations, probably due to high mutation rates of viral RNA polymerase by several virus passages. Even though the current JEV vaccine strains have been maintained and produced for a long period of time, the genetic characterization of them have been rarely changed. However, since the mid 1990's, molecular epidemiology of JEV has been changed sharply from genotype 3 to genotype 1 in Korea, further studies on new vaccine strains to genotype 1 is required for more effective prevention in the field.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.148.2-149
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• 2003

Cucumber fruit mottle mosaic tobamovirus (CFMMV) causes severe mosaic symptoms with yellow mottling on leaves and fruits, and occasionally severe wilting of cucumber plants. No genetic source of resistance against this virus has been identified. The genes coding for the coat protein or the putative 54-kDa replicase were cloned into binary vectors under control of the SVBV promoter. Agrobacterium-mediated transformation was peformed on cotyledon explants of a parthenocarpic cucumber cultivar with superior competence for transformation. R1 seedlings were evaluated for resistance to CFMMV infection by lack of symptom expression, back inoculation on an alternative host and ELISA. From a total of 14 replicase-containing R1 lines, 8 exhibited immunity, while only 3 resistant lines were found among a total of 9 CP-containing lines. Line 144 homozygous for the 54-kDa replicase was selected for further resistance analysis. Line 144 was immune to CFMMV infection by mechanical and graft inoculation, or by root infection following planting in CFMMV-contaminated soil. Additionally, line 144 showed delay of symptom appearance following infection by other cucurbit-infecting tobamoviruses. Infection of line 144 plants with various potyviruses and cucumber mosaic cucumovirus did not break the resistance to CFMMV. The mechanism of resistance of line 144 appears to be RNA-mediated, however the means is apparently different from the gene silencing phenomenon. Homozygote line 144 cucumber as rootstock demonstrated for the first time protection of a non-transformed scion from soil inoculation with a soil borne pathogen, CFMMV.

• Research in Plant Disease
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• v.28 no.4
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• pp.231-236
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• 2022

Oxypetalum coeruleum, commonly known as Tweedia, is a perennial herbaceous plant of the Apocynaceae family native to southern Brazil and Uruguay. Tweedia plants are grown as one of the most popular ornamental flowers for floral arrangement in Korea. In May 2021, several tweedia plants in a single greenhouse in Gimje, Jeollabuk-do were found to show virus-like symptoms including necrotic rings, vein-clearing, chlorotic mottle, and mosaic on the leaves, and necrosis on the stems. Here, we have identified tomato spotted wilt virus (TSWV) in symptomatic tweedia leaves by applying high-throughput RNA sequencing. In the result, a single infection by TSWV was verified without mixed infections of different virus species. To confirm the presence of TSWV, a reverse transcription polymerase chain reaction was performed with a specific primer set to the N gene of TSWV. The complete genomic sequence of L, M, and S segments of TSWV 'Oxy' isolate were determined and deposited in GenBank under accession numbers LC671525, LC671638, and LC671639, respectively. In the phylogenetic tree analysis by maximum likelihood method, 'Oxy' isolate showed a high relationship with TSWV 'Gumi' isolate from Gerbera jamesonii in Gyeongsangbuk-do, Korea; for all three RNA segments. To our knowledge, this is the first report of TSWV infection of O. coeruleum in Korea.

• Applied Biological Chemistry
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• v.38 no.3
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• pp.224-231
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• 1995

To increase the expression of a foreign protein in transgenic plant, the benefits of 5'-untranslated leader sequences of tobacco mosaic virus (TMV) RNA or soybean glycinin gene, Gy2, fused to a protein coding sequence were exploited. pGA643-derived plasmid contains 355 promoter of cauliflower mosaic virus, protein coding sequence of maize 10 kDa zein (10kZ) and Gy2 terminator. The leader from Gy2 or TMV RNA was inserted between the promoter and the coding sequence in each construct. The recombinant DNAs were introduced into tobacco plants by Agrobacterium mediated leaf disc transformation method. Although the transgene without the leader had more transcripts than the others, mRNAs containing the leader were translated more efficiently. It might be due to difference in the length of 5'-untranslated sequence and context surrounding the AUG codon, but could be sequence specific rather. These results suggest that the leader sequences of Gy2 and TMV play important roles as an enhancer in translational control of foreign gene in transgenic tobacco plant.