• 미생물학회지
• /
• 제29권4호
• /
• pp.221-225
• /
• 1991

The RNA transcripts produced from in vitro transcription reaction of BTV core were analyzed on agarose-urea gel. Fast migrating abortive RNAs, in addition to full length species of RNA, were observed. Fast migrating RNAs extracted from agarose-urea gel were hybridized to all 10 segments of genomic ds RNA, while solw migrating RNAs extracted from agarose-urea gel were hybridized only to the large and medium size genomic ds RNA. These results indicate that fast migrating RNA transcripts are most likely the products of abortive transcription.

• Journal of Microbiology
• /
• 제34권1호
• /
• pp.61-67
• /
• 1996

Double-stranded RNA (dsRNA) viruses and single-stranded RNA(ssRNA) viruses were detected in a strain of Pleurotus mushroom cultivated in a farm. Those fungal virsus were purified in the pH 6.0 or pH 7.2 using CsCI or Cs$_{2}$SO$_{4}$ buoyant density centrifugation. Each viral particles were not completely separated at any trials. However, mushroom bacili-form virus contains a single major nucleic acid with 0.7 Kb ssRNA, which might code for 20 Kd viral capsid protein. The dsRNAs are encapsidatred into spherical-form viruses, whereas ssRNA viral genomes are encapsidated into two different sizes of bacili-form particles. A healthy-looking mushroom also contained some spherical-form viruses with dsRNAs. Laboratory strains of Pleurotus ostreatus and a cultivated strain of P. sajor-caju did not show any viral particles. Mushrooms with specific disease symptoms. however, contained at least four different sizes of spherical-form viruses. Thus, we concluded that a bacilli-form virus case a severe disease symptoms of adnormal on mushroom development.

• The Plant Pathology Journal
• /
• 제19권3호
• /
• pp.152-158
• /
• 2003

Stromata of the white root rot fungus, Rosellinia necatrix, were produced on diseased roots although they were reported to develop rarely in nature. Forty-two (42) out of 47 samples produced synnemata while 23 developed stromata. Forty-seven (47) isolates obtained from diseased root samples were divided into 24 mycelium compatibility groups (MCGs). Sixteen (16) out of 24 MCGs produced stromata. Single ascospore isolates from 10 stroma samples produced dsRNA-containing isolates from diseased tissue beneath stromata. The frequency of synnema production on axenic culture varied among isolates with different origin. The dsRNA was not transmitted vertically to the ascospore offspring despite the infection of various dsRNA in the parental isolates. The dsRNA was absent in 35 ascospore isolates in two stroma samples that originated from the isolates, in which dsRNA was not eliminated by hyphal tip isolation. Consequently, sexual reproduction in the white root rot fungus was suggested to produce propagules as a new infection source and to have the function to eliminate infectious factors such as mycoviruses.

• 한국식물병리학회:학술대회논문집
• /
• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
• /
• pp.150.3-151
• /
• 2003

To analyze the genome of Cucumber mosaic virus(CMV) in pepper, we developed a new extraction method for double-stranded RNA(dsRNA). To isolate the dsRNA, 0.1g of pepper leaves homogenized with 1ml of 5${\times}$EXB extraction buffer[0.5M glycin, 0.5M NaCl, 5mM EDTA(pH9.0/NaOH), 10％ Sodium N-lauryl salcosinate(NLS), 10％ Sodium dodecylsulfate(SDS)] and purified with the 1/4 volume of phenol： chloroform： isoamylalcohol(25：24：1). dsRNAs from the aqueous phase was precipitated with isopropanol. This procedure was able to detect a minimal amount of dsRNA from CMV infected plant tissue and to distinguish different CMV satellite RNAs by polyacrylamide gel electrophoresis(PAGE). Moreover, this method can be applied CMV infected in pepper or Rice dwarf virus (RDV) infected rice.

• 미생물과산업
• /
• 제16권2호
• /
• pp.10-13
• /
• 1990

Fungal Virus에 대한 연구는 60년대말 double-stranded RNA(dsRNA)가 interferon의 생성을 촉진시킨다는 보고가 나온 이후 dsRNA를 많이 얻을수 있는 재료로써 이 virus에 대한 연구가 본격적으로 진행되었다. 현재까지 60여종 이상의 genus와 200가지 이상의 species에서 virus의 존재가 분리 또는 확인되어 있다. 여기서는 Ustilago maydis의 killer activity를 유발하는 virus에 대한 일반적인 특징과 이 virus의 transcription 과정에 대해 설명하고자 한다.

• The Plant Pathology Journal
• /
• 제20권3호
• /
• pp.200-205
• /
• 2004

The partial nucleotide sequences of the genomic dsRNA mycovirus infecting Pleurotus ostreatus isolates ASI2223 and Suhan were determined and compared with those of mycoviruses belonging to partitiviruses and totiviruses. Partial nucleotide sequences of the purified dsRNA from ASI2223 and Suhan showed RNA-dependent RNA polymerase sequences that are closely related to those of partitiviruses, including Fusarium poae virus 1, Fusarium solani virus, Rhizoctoniasolani virus, Discula destructiva virus 2, and Oyster mushroom isometric virus 2. Specific primers were designed for RT-PCR detection of dsRNA viruses from the P. ostreatus isolate ASI2223 and Suhan. Two virus specific primer sets were found to specifically detect each virus among six sets of designed oligonucleotide primers. Collectively, these results suggest that dsRNA mycoviruses from P. ostreatus isolates ASI2223 and Suhan belong to the family Partitiviridae, although, they are not the same virus species. Our results also suggest that these virus-specific primer sets can be employed for the specific detection of each viral sequence in infected tissues.

• 식물병연구
• /
• 제14권3호
• /
• pp.223-225
• /
• 2008

2005년 전북 고참에서 채집한 옥수수 이병주로부터 벼검은줄오갈병 바이러스를 동정하였다. 이들 이병주로부터 게놈 dsRNA를 추출하여 polyacrylamide gel 전기영동으로 게놈 패턴을 분석 하였다. 전기영동 결과 이미 알려진 10개의 분절게놈을 확인하였으며 채집지역별 isolate에서 게놈 dsRNA이동도의 차이를 확인하였다. 추출된 dsRNA를 주형으로 하여 S10의 full-length 특이 primer를 사용하여 RT-PCR한 결과 1,801의 예상되는 band를 확인하여 RBSDV로 동정하였다. S10을 pGEM-T vector에 크로닝하여 염기서열 분석 결과 1,801nt, 559aa로 구성되어 있었다. 이는 벼에 발생하는 RBSDV S10의 크기와 동일하였으며 상동성 분석결과 18개 염기에서 변이가 확인되어 99%의 상동성을 나타내었다.

• Mycobiology
• /
• 제34권2호
• /
• pp.61-66
• /
• 2006

Chestnut blight disease caused by Cryphonectria parasitica is widely distributed throughout chestnut tree plantations in Korea. We surveyed 65 sites located at 9 provinces in South Korea, and isolated 248 virulent and 3 hypovirulent strains of chestnut blight fungus. Hypovirulent strains had dsRNA virus in the cytoplasm, which is one of the typical characteristics of hypovirulent strains. In addition, they showed more characteristics of hypovirulent strains, i.e., suppressed conidiation, reduced pigmentation in colony color, and reduced phenol oxidase activity as well as reduced pathogenicity. Hypovirulent strains, KCPH-22, KCPH-135 and KCPH-136, had a genomic dsRNA band with the molecular weight of 12.7 kb, which is the L-dsRNA of CHV1. They also had a 2.7 kb defective dsRNA band. Single conidia isolated from hypovirulent strains were cultured and various phenotypes and absence of dsRNA bands were obtained from single conidial cultures, which means that hypovirulence transmission is unstable in asexual reproduction and variations in viral heredity by asexual reproduction. Biocontrol trial using hypovirulent strains was also carried out in the chestnut tree plantations, and canker expansion in the treated trees was stopped and healed by callus formation at the margin of the canker. These results show the potentials in successful biocontrol of chestnut blight if the vegetatively compatible hypovirulent strains could be directly used around the canker formed by compatible virulent strains.

• Fisheries and Aquatic Sciences
• /
• 제17권3호
• /
• pp.377-380
• /
• 2014

RNA interference (RNAi)-mediated transcriptional knock-down of Crassostrea gigas big defensin 1 and 2 genes (Cg-BigDef1 and Cg-BigDef2) was investigated. The cDNA sequences of Cg-BigDef1 and Cg-BigDef2 were identical, excluding an additional fragment of 20 nucleotides in Cg-BigDef1; thus, a long double-stranded RNA (dsRNA) targeting the mRNA of Cg-BigDef2 effectively downregulated both Cg-BigDef2 and Cg-BigDef1. In addition, long dsRNA targeting green fluorescent protein (GFP) did not affect transcription of the two big defensin genes. These results suggest that the transcriptional downregulation of Cg-BigDef1 and Cg-BigDef2 was mediated by sequence-specific RNA interference (RNAi). Despite injection of long dsRNA targeting Cg-BigDef2 into only the adductor muscle, knock-down of Cg-BigDef1 and Cg-BigDef2 was observed in the adductor muscle, hemocytes, mantle, and gills, suggestive of systemic spread of RNAi in C. gigas. Furthermore, the inhibitory effect of dsRNA persisted until 72 h post-injection, indicative of a long-lasting RNAi-mediated knock-down of target genes.

• 한국식물병리학회지
• /
• 제11권2호
• /
• pp.173-178
• /
• 1995

동물계에서 항바이러스와관련된 dsRNA 의존성 인산화 효소(PKR)의 유전자를 식물체에서 발현시킬 경우 PKR에 의한 단백질합성 및 식물바이러스의 증식조절 가능성에 대한 기초자료를 확보하기 위하여 사람에서 분리된 PKR cDNA를 Agrobacterium 방법에 의하여 연초식물체(Nicotiana tabacum cv. Xanthi-nc)로 형질전환시켰다. HindIII/PstI처리에 의해 얻어지는 약 1.8kb의 phPKR cDNA절편을 일련의 유전자 조작 방법을 통하여 식물발현벡터인 pBI121에 도입하여, p12168을 재조합하였다. 이를 A. tumefaciens LBA 4404에 형질전환시켜 연초식물체형질 전환에 이용하였다. 2mg/l BA와 0.5mg/l NAA가 포함되고 100$\mu\textrm{g}$/ml의 kanamycin이 첨가된 MS배지에서 shooting시킨 후 phytohormone이 첨가되지 않은 MS배지상에서 rooting을 시켜 형질전환 연초식물체를 얻었으며, 형질전환식물체는 정상식물체와 유사한 생육양상을 나타내었다. 형질전환식물체의 유전자도입은 hPKR cDNA의 전사부여는 RT-PCR 방법에 의하여 확인되었다.