• 미생물학회지
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• 제29권1호
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• pp.34-39
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• 1991

The BTV core associated transcriptase activity, assayed by acid precipitable counts, was reduced to an undetectable level after treat the core with .$100{\mu}{\M}$ CDDP. When the RNA transcripts prepared from the CDDP treated BTV core were analysed on agaroseurea gel, it was observed that the band intensity of the large size RNA was reduced while the band intensity of the small size RNA was enhanced. Northern blot analysis showed that much of the small size RNAs appeared to be prematurely terminated transcripts. These results suggest that CDDP adduction to the template RNA blocks chain elongation process of the virion bound transcriptase that is ultimately responsible for the inactivation of BTV infectivity.

• 미생물학회지
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• 제26권3호
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• pp.149-154
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• 1988

Japanese Encephalitis Virus(JEV) can be detected conveniently by the use of biotinylated cDNA probe. To prepare biotinylated probe aminoallyl dUTP was first synthesized chemically to reverse transcribe the virial RNA. The allylamine-labeled cDNA was then converted to the biotin-cDNA by the reaction with an activated biotin ester, NHS-ACA-biotin. The JEV genomic RNA was hybridized to the biotinylated cDNA probe on nitrocellulose filter and visualized colorimetrically by streptavidin complexes with alkaline phosphatase polymer. Sensitivity of the detection system was determined by estimating the amount of the JEV genomic RNA through comparison with signals generated from the biotinylated and $^{32/P}$ -labeled probes. It was found that the biotin probe was as sensitive as $^{32/P}$ -cDNA probe which can detect 50pgs of the target RNA.

• The Plant Pathology Journal
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• 제39권6호
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• pp.592-599
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• 2023

A defective RNA3 (D3Yα) of strain Y of cucumber mosaic virus (CMV-Y) was examined on host-specific maintenance, experimental conditions, and a viral factor required for its generation in plants. D3Yα was stably maintained in cucumber but not in tomato plants for 28 days post inoculation (dpi). D3Yα was generated in Nicotiana tabacum or N. benthamiana after prolonged infection in the second and the third passages, but not in plants of N. benthamiana grown at low temperature at 28 dpi or infected with CMV-Y mutant that had the 2b gene deleted. Collectively, we suggest that generation and retention of D3Yα depends on potential host plants and experimental conditions, and that the 2b protein has a role for facilitation of generation of D3Yα.

• The Plant Pathology Journal
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• 제25권2호
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• pp.142-150
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• 2009

The complete genome sequences of RNA3 and RNA4 of the 13 different Rice stripe virus (RSV) isolates were determined and characterized in this study to address the possible causes of the recent re-emergence of RSV that affected many rice fields in Korea. The genome size of each RNA segment varied among isolates and significant differences were observed in the intergenic region. There was up to 4% average divergence in the RNA4 nucleotide sequence among 13 Korean isolates and only 1.4% in the RNA3. Phylogenetic relationships among different Korean isolates revealed that there were at least 2 types of RNA3 and 4 distinct types of RNA4 genomes present in Korea. However, Korean isolates with one type of RNA3 predominate over the other while the occurrences of the RSV Korean isolates with the 4 types of RNA4 genome were not correlated to specific geographical areas. Results further indicate that RNA4 had diverged more than RNA3 and these differences in accumulation of mutations in the individual RNA segments indicate that genetic reassortment were likely to contribute to the genetic divergence in the 13 Korean isolates. All of the Korean-RNA3 sequences except for one isolate grouped with Chinese isolates (JY and Z). In contrast, the RNA 4 sequences segregated together with either Chinese (JY and Z) and Japanese (M and T) isolates but genetic relationships of Korean isolates- RNAs 3 and 4 segments to Chinese-Y isolate were low. Altogether, these results suggest that the occurrence of mixtures of RNAs 3 and 4 genotypes in the natural population of RSV may have contributed to the sudden outbreak in Korea.

• 한국식물병리학회지
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• 제12권2호
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• pp.176-181
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• 1996

Aster yomena로부터 분리한 오이 모자이크 바이러스(cucumber mosaic virus) (CMV-As)의 RNA4로부터 완전한 길이의 cDNA를 합성하고 그 전체적인 염기서열(1,043 nt`s)을 결정하였다. CMV-As RNA4는 73개의 염기로 구성된 5`말단의 leader 부위, 657개의 염기로 구성된 외피단백질(coat protein) 유전자 부위 및 312개의 염기로 구성된 3` 말단의 비번역 부위로 구성되어 있음을 확인하였다. 외피단백질 유전자 부위의 염기서열을 다른 계통의 CMV와 비교해 볼 때 그 염기서열이 보전적으로 존재하고 있으나 그 외의 부분은 다양함을 확인하였다. 특히 3` 말단부위의 61개의 염기로 구성된 부위(959-1019)는 다른 계통의 CMV에서는 상당히 유사하지만 CMV-As도 다른 CMV처럼 tRNA와 유사한 구조를 역시 형성함을 확인하였다. CMV-As의 RNA4 염기서열을 다른 계통의 CMV와 비교할 때 CMV-I17F와 가장 유사하였으며(91.9%) S형의 CMV-M과는 가장 낮은 동일성을 보였다(71.1%). 외와 같은 염기성열의 비교 결과와 EcoRI 제한효소 인식부위의 존재로 미루어 CMV-As는 WT형으로 분류된다.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2003년도 International Meeting of the Microbiological Society of Korea
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• pp.83-86
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• 2003

Major advances in positive-sense RNA virus research have been facilitated by the development of reverse genetics systems. These systems consist of an infectious cDNA clone that encompasses the genome of the virus in question. This clone is then used as a template for the subsequent synthesis of infectious RNA for the generation of synthetic viruses. However, the construction of infectious cDNA for the Japanese encephalitis virus (JEV) has been repeatedly thwarted by the instability of its cDNA. As JEV is an important human pathogen that causes permanent neuropsychiatric sequelae and even fatal disease, a reliable reverse genetics system for this virus is highly desirable. The availability of this tool would greatly and the development of effective vaccines as well as facilitate studies into the basic biology of the virus, including the molecular mechanisms of viral replication, neurovirulence, and pathogenesis. We have successfully constructed a genetically stable infectious JEV cDNA containing full-length viral RNA genome. Synthetic RNA transcripts generated in vitro from the cDNA were highly infectious upon transfection into susceptible cells, and the cDNA remained stable after it had been propagated in E. coli for 180 generations. Using this infectious JEV cDNA, we have successfully expressed a variety of reporter genes from the full-length genomic and various subgenomic RNAs in vitro transcribed from functional JEV cDNAS. In summary, we have developed a reverse genetics system for JEV that will greatly facilitate the research on this virus in a variety of different fields. It will also be useful as a heterologous gene expression vector and aid the development of a vaccine against JEV.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2000년도 추계 학술발표회
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• pp.57.1-57
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• 2000

• Bulletin of the Korean Chemical Society
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• 제31권4호
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• pp.1021-1024
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• 2010

• The Plant Pathology Journal
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• 제28권1호
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• pp.75-80
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• 2012

$Potato$ $virus$ $X$ (PVX) contains $cis$-acting elements including stem-loop 1 (SL1) RNA at the 5' region; SL1 is conserved among all potexviruses. The SL1 at the positive-sense RNA, SL1(+), is required for PVX RNA replication, cell-to-cell movement, and translation. Previous research demonstrated that SL1(+) RNA also serves as the origin of assembly for encapsidation of PVX RNA. To identify the essential sequences and/or regions for capsid protein (CP) subunit recognition within SL1(+) RNA, we used electrophoretic mobility shift assays (EMSA), UV cross-linking, and yeast three-hybrid analyses. The EMSA and UV cross-linking analyses with PVX CP subunits and RNA transcripts corresponding to the SL1(+) RNA showed that the SL1(+) RNA formed complexes with CP subunits. We also conducted EMSA and yeast three-hybrid analyses with RNAs containing various mutations of SL1(+) RNA elements. These analyses indicated that SL1(+) RNA is required for the interaction with PVX CP and that the RNA sequences located at the loop C and tetra loop of the SL1(+) are crucial for CP binding. These results indicate that, in addition to being important for RNA accumulation, the SL1(+) RNA from the 5' region of the PVX genome is also required for specific binding of PVX CP.

• 식물병연구
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• 제7권3호
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• pp.155-163
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• 2001

기주반응 실험을 통하여 무병징 또는 엷은 병징을 발현하는 satRNA 보유 Paf-CMV 및 Rs2-CMV를 고추 CMV병의 방제를 위한 약독바이러스로 공시하여 교차방어효과를 검정하였다. 공시한 satRNA-CMV는 모두 agar gel diffusion test에서 강독계로 공시한 Mf-CMV의 항원과 융합하는 침강선을 나타내 subgroup I의 혈청 형으로 판단되었다. 이들 약독계 satRNA-CMV의 물리적성질은 내희석성이 $10^{-4}$으로, 강독계 Mf-CMV나 satRNA를 보유하고 있는 Ap-CMV보다 낮게 나타났으나, 내열성 및 내보존성 은 차이를 보이지 않았다. 공시한 satRNA의 염기서열을 결정한 결과, Rs2-satRNA는 335염기, Ap-satRNA 347염기, Paf-satNA 386염기로 구성되어 있었다. 이들 염기서열을 이미 보고된 Y-CMV의 satRNA와 비교한 결과, 양 말단 영역 특히 5'말단으로부터 80염기 및 3'말단으로부터 174염기는 안정된 conserved sequence를 나타냈다. 그러나 중간영역의 염기서열에서는 .많은 변이를 나타냈고, 특히 병징과 관련된 domain으로 보고된 영역에서 Paf-satRNA의 염기서열은 다른 계통의 satRNA에 비하여 많은 차이를 보였다. 각 satRNA의 cDNA로부터 전사시킨 transcript RNA를 Mf-CMV의 게놈RNA와 혼합하여 고추에 접종한 결과, 본래의 각 satRNA-CMV를 접종하였을 때와 마찬가지로 Paf-satRNA 및 Rs2-satRNA의 transcript 와 혼합한 Mf-CMV에 감염된 고추의 병징이 약하게 발현되었다. 선발된 약독CMV의 강독계 바이러스에 대한 교차방어효과를 검정하기 위하여 Paf-CMV 및 Rs2-CMV를 접종한 고추와 담배에 강독 Mf-CMV를 challenge한 후 교차방어 지속효과를 검정한 결과, Paf-CMV를 접종한 고추와 담배는 모두 Mf-CMV를 challenge 접종한 3주후까지 병징이 발현되지 않았으나, Rs2-CMV를 접종한 식물은 challenge 2주 후에 약 반수의 개체에서 강독계 병징이 발현되었다. 또한 challenge바이러스의 농도별 교차방어효과에서도 Paf-CMV를 접종한 고추에 정제한 Mf-CMV를 0.2 mg/ml 이하의 농도로 challenge한 경우, 접종 30일이 되었을 때까지 병징이 발현되지 않았으나, Rs2-CMV에서는 일부의 개체에서 병징이 발현되어 강독계에 대한 교차방어의 효과가 일정하게 나타나지 않았다. 한편 고추의 유묘에 약독CMV를 접종한 다음, 포장에 재배하면서 바이러스병징이 발현되는 개체를 조사한 결과, 약독 CMV접종 30-60일 후에 Paf-CMV 접종구에서 1.8-6.4%, Rs2-CMV 접종구에서 8.2-18.3%그리고 무접종구에서 2.7-47.2%의 바이러스병징이 발현됨으로서 Paf-CMV의 강독계 CMV의 감염에 대한 교차방어효과가 안정되고 높게 나타났다..