• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.138.2-139
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• 2003

Grapevine leafroll-associated 1 virus (GLRaV-1) and Grapevine leafroll-associated 3 virus (GLRaV-3), member of the genus Ampelovirus, are important viral disease of grapevine in the world. these viruses transmitted only dicotyledonous host by vectors such as mealybugs and there is no suitable herbaceous host for virus. The diseased leaves turn yellowish or reddish depending on cultivars and viruses. Viruses are existed at low concentration and ununiformly distribution in grapevine. Using small-scale double-stranded RNA (dsRNA) extraction method, reverse transcription and polymerase chain reaction (RT-PCR) product of 1Kb long which encoded of coat protein (CP) gene for both viruses was successfully amplified with a specific primers. The RT-PCR product was cloned into the plasmid vector and its nucleotide sequences were determined from selected recombinant cDNA clones. Sequence analysis revealed that the CP of GLRaV-1 consisted of 969 nucleotide, which encoded 323 amino acid residues and CP of GLRaV-3 consisted of 942 nucleotide, which encoded 314 amino acid residues. The CP of GLRaV-1 and GLRaV-3 has 93.8％ and 98.7％ amino acid sequence identities, respectively.

• KSBB Journal
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• v.31 no.4
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• pp.300-311
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• 2016

Recently gene delivery has been designed newly using bioactive biomaterial and applied in the various field by many researchers. In this study, we proposed a new gene delivery system which has the capability of targeting effect in the specific tissue and remarkably enhanced transfection efficiency. We investigated $^1H-NMR$ spectroscopy, particle size analyzer and gel retardation to confirm the correct preparation of gene delivery. Also, we identified the hemo-compatibility of gene delivery by hemolysis assay, non-cytotoxicity by MTT test and transfection efficiency. The uptake mechanism of the gene carrier was confirmed using inhibitor agent such as sodium azide, indomethacin, quercetin, colchicine, and chloropromazine. As a results, it was identified that gene carrier prepared by in this study entered in the cell by the microtubule-dependent, energy-dependent and clathrin-mediated endocytosis pathway.

• Molecules and Cells
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• v.37 no.7
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• pp.518-525
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• 2014

Gammaherpesvirus (${\gamma}HV$) infection of the central nervous system (CNS) has been implicated in diverse neurological diseases, and murine ${\gamma}HV$-68 (MHV-68) is known to persist in the brain after cerebral infection. The underlying molecular mechanisms of persistency of virus in the brain are poorly understood. Here, we characterized a unique pattern of MHV-68 persistent infection in neuroblastoma cells. On infection with MHV-68, both murine and human neuroblastoma cells expressed viral lytic proteins and produced virions. However, the infected cells survived productive infection and could be cultured for multiple passages without affecting their cellular growth. Latent infection as well as productive replication was established in these prolonged cultures, and lytic replication was further increased by treatment with lytic inducers. Our results provide a novel system to study persistent infection of ${\gamma}HVs$ in vitro following de novo infection and suggest application of MHV-68 as a potential gene transfer vector to the brain.

• The Journal of Korean Society of Virology
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• v.28 no.1
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• pp.21-30
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• 1998

Synthetic genes encoding the gag p24 and the part of the envelope protein gp41 of the human immunodeficiency virus (HIV-1) were cloned and overexpressed as fusion proteins in Escherichia coli, using an expression vector carrying T7 promoter and the poly-histidine leader, sequence. The overexpressed p24 fusion protein was purified by centrifugation, Ni-affinity chromatography and CM-sepharose chromatography. The overexpressed gp41 fusion protein was purified by centrifugation, $C_4$ chromatography and DEAE-sepharose chromatography. The purified fusion proteins showed a high level of purity and immunoreactivity in SDS-polyacrylamide gel electrophoresis and western blot analysis. These results suggest that this prokaryotic - expression purification method is suitable for obtaining a large amount of the viral antigen which may be useful for screening of antibodies to HIV-1 in human blood samples.

• BMB Reports
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• v.34 no.4
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• pp.371-378
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• 2001

The Glycoprotein D (gD) gene of the HSV-1 strain F was cloned, sequenced, recombinated into the HcNPV (Hyphantria cunea nuclear polyhedrosis virus) expression vector and expressed in insect cells. The gD gene was located in the 6.43 kb BamHI fragment of the strainF. The open reading frame (ORF) of the gD gene was 1,185 by and codes 394 amino acid residues. Recombinant baculoviruses, GD-HcNPVs, expressing the gD protein were constructed. Spodoptera frugiperda cells, infected with the recombinant virus, synthesized a matured gX-gD fusion protein with an approximate molecular weight of 54 kDa and secreted the gD proteins into the culture media by an immunoprecipitation assay The fusion gD protein was localized on the membrane of the insect cells, seen by using an immunofluorescence assay The deduced amino acid sequence presents additional characteristics compatible with the structure of a viral glycoprotein: signal peptide, putative glycosylation sites and a long C-terminal transmembrane sequence. These results indicate the utility of the HcNPV-insect cell system for producing and characterizing eukaryotic proteins.

• International Journal of Industrial Entomology and Biomaterials
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• v.21 no.2
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• pp.157-162
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• 2010

Open reading frame 35 (bm35) of the Bombyx mori nucleopolyhedrovirus (BmNPV) is a special gene whose homologues are only found in some group-I nucleopolyhedroviruses, suggesting that bm35 plays a specific role in the viral life cycle. This paper described the characterization of BmNPV bm35. Computerassisted sequence analysis shows that a putative RING finger motif is observed in the protein, Bm35 encoded by bm35. The coding sequence of bm35 was amplified and subcloned into the vector pET30a(+) and the $(His)_6$-tagged fusion protein His-Bm35 was expressed in the Escherichia coli BL21 (DE3) LysS cells. The bm35 transcript and Bm35 protein were detected in BmNPV-infected BmN cells at 12~48 h post infection (p.i.) by RT-PCR and Western blot analysis using the polyclonal antibody generated by immunizing a rabbit with purified $(His)_6$-tagged Bm35, suggesting that bm35 is synthesized in the late stage of BmNPV infection cycle. Bm35 was not a structural component associated with budded virus (BV) and occlusion derived virus (ODV). These data indicated that bm35 is a functional gene in the BmNPV life cycle.

• Korean Journal of Veterinary Research
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• v.31 no.3
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• pp.311-318
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• 1991

The DNA fragment representing for Pseudorabies gp50 and gp60(Shope) was cloned by recombinant techniques. The viral DNA was extracted from the infected cells and digested with Bam HI. The 6.8 Kb of Bam HI fragment was isolated from agarose gel and further digested with Nde I followed by Klenow treatment. The blunt ended 4.9Kb fragment was cloned into pTZ18R plasmid vector. The upstream region of gp50 was further manipulated to remove its 5' promoter region and create EcoRl site for possible eukaryotic expression system. The result of partial sequencing of cloned DNA indicated that Shope strain showed 95% homology with gp50 of Rice strain.

• Journal of Microbiology and Biotechnology
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• v.22 no.6
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• pp.866-871
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• 2012

A novel cholesterol-based cationic lipid containing a tri-2-hydroxyethylamine head group and ether linker (Chol-THEA) was synthesized and examined as a potent gene delivery vehicle. In the preparation of cationic liposome, the addition of DOPE as helper lipid significantly increased the transfection efficiency. To find the optimum transfection efficiency, we screened various weight ratios of DOPE and liposome/DNA (N/P). The best transfection efficiency was found at the Chol-THEA:DOPE weight ratio of 1:1 and N/P weight ratio of 10~15. Most of the plasmid DNA was retarded by this liposome at the optimum N/P weight ratio of 10. The transfection efficiency of Chol-THEA liposome was compared with DOTAP, Lipofectamine, and DMRIE-C using the luciferase assay and GFP expression. Chol-THEA liposome with low toxicity had better or similar potency of gene delivery compared with commercial liposomes in COS-7, Huh-7, and MCF-7 cells. Therefore, Chol-THEA could be a useful non-viral vector for gene delivery.

• Korean Journal of Agricultural Science
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• v.44 no.3
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• pp.348-358
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• 2017

The green peach aphid, Myzus persicae, is one of the most serious insect pest and a vector for a multitude of viral diseases to many crops, vegetables, ornamentals, and fruit trees in the world. A large number of aphids can reduce plant vigor and cause defoliation. Many insecticides have been developed and applied to control the green peach aphid. However, this aphid has displayed a remarkable ability to establish resistance to almost every insecticide. We treated 5 different insecticides registered for M. persicae on pepper leaves and investigated the effects of the insecticides by measuring the time it took to achieve 90% control of the aphids. Acetamiprid worked faster than any other insecticides while cyantraniliprole showed the slowest insecticidal effect. Pymetrozine, pyrifluquinazon, and spirotetramet provided 90% control within similar time. Iwol population's control value was higher than any other populations 24 hours after treatment. When five different unregistered insecticides for M. persicae were treated on pepper leaves, no insecticidal effect was found for gamma-cyhalothrin and novaluron and spinosad showed an insecticidal effect of up to 70% in Iwol population only. Although chlorfenapyr and dinotefuran were not registered for M. persicae, their insecticidal effects were found to be 90% or higher.

• Bulletin of the Korean Chemical Society
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• v.22 no.1
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• pp.46-52
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• 2001

To evaluate the non-ionic polymer, poly(ethylene glycol) (PEG), as a component in cationic copolymers for non-viral gene delivery systems, PEG was coupled to polyethylenimine (PEI). We present the effects of different degrees and shapes of pegylation of PEI on cytotoxicity, water solubility and transfection efficiency. This work reports the synthesis and characterization of a series of cationic copolymers on the basis of the conjugates of PEI with PEG. The modified molecules were significantly less toxic than the original polymer. Moreover, the chemical modification led to enhancement of their solubility. The comparison of pegylated PEIs with different degrees of derivation showed that all the polymers tested reached comparable levels of transgene expression to that of native PEI. As assessed by agarose gel electrophoresis, even highly substituted PEI derivatives were still able to form polyionic complexes with DNA. However, aside from an increase in solubility and retention of the ability to condense DNA, methoxy-PEG-modified PEIs resulted in a significant decrease in the transfection activity of the DNA complexes. In fact, the efficiency of the copolymer was compromised even at a low degree of modification suggesting that the PEG action resulting from its shape is important for efficient gene transfer. The mode of PEG grafting and the degree of modification influenced the transfection efficiency of PEI.