• 한국식물병리학회지
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• 제14권6호
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• pp.636-641
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• 1998

Digoxigenin (DIG) was used to prepare nucleic acid probe for the detection of RNA of potato leafroll virus (PLRV) in the potato leaf extracts. The 0.6 kb coat protein (CP) gene cDNA of PLRV in plasmid pSPT 18 vector was labeled with digoxigenin by in vitro run-off transcription and then used for cRNA probe. In the several buffers tested for increase the total RNA extraction efficiency AMES buffer was the most suitable for this detection method. The RNA extracts from potato leaves shown symptoms of PLRV were dot blotted onto nylon membrane and hybridized with labeled RNA probes. After hybridization, labeled RNA bound to PLRV RNA on membrane was detected with anti-digoxigenin alkaline phosphatase. 5-bromo-4-chloro-3-indolyl-phosphate/nitroblue tetrazolium (NBT) salt and CSPD were used as substrate for colorimetric and film exposure detection, respectively. These detection methods were very sensitive allowing for detection of 1/32 diluted total RNA extract from 100 mg leaf tissue.

• 한국응용곤충학회지
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• 제49권1호
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• pp.23-29
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• 2010

토마토나 고추 등의 유용작물에 자주 발생하여 생산량을 저하시키는 오이모자이크병에 대한 저항성을 높여 생산량을 증대시키기 위하여 개발된 CMVP0-CP (Cucumber mosaic virus-coat protein) gene이 삽입된 유전자 변형 CMVP0-CP 고추 (line 7)에 대한 환경위해성을 평가하였다. 2007년 고추의 생육기간 동안 절지동물의 군집구조를 3회(6월 19일, 7월30일, 8월 31일)에 걸쳐 조사를 하였다. 두 가시의 고추, 즉 모본(P915, nTR)고추와 유전자변형 고추(CMVP0-CP (line 7), TR)의 꽃과 잎에 서식하는 곤충을 포함한 절지동물의 군집구조를 파악하기 위하여 곤충을 포획할 수 있는 곤충진공포획기를 사용하여 절지동물을 정량적으로 채집하였다. 절지동물의 군집은 채집 시기별로 출현 종과 빈도수에 차이는 있었지만 두 작물간의 군집 구조는 통계학적으로 차이를 나타내지 않았다. 지금까지 수행된 본 연구결과를 근거로 판단하여 볼 때, 유전자 변형 고추로 인한 비표적 생물체의 군집구조가 모본 고추의 군집구조와 차이가 나타나지 않아 환경 위해서도 없는 것으로 볼 수 있지만, 좀더 확실한 유전자변형생물체인 고추의 환경위해성이 없다는 결론을 얻기 위해서는 추가적인 연구가 수행되어져야 할 것이다.

• The Plant Pathology Journal
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• 제20권3호
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• pp.212-219
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• 2004

Three isolates of Cucumber mosaic virus (CMV) from lily plants showing mosaic and distortion symptoms were detected by reverse-transcriptase polymerase chain reaction (RT-PCR) using primers specific to Cucumovirus genus namely, LK-CMV, LK4-CMV, and LKS-CMV. Restriction enzymes patterns of the RT-PCR products revealed that the lily isolates belonged to subgroup IA of CMV. In terms of biological properties, the lily isolates have highly similar but distinct pathogenicity as reported in other lily strains and ordinary strains of CMV. To characterize the molecular properties, cDNAs containing coat protein (CP) gene and 3' non-coding region (NCR) of RNA3 for the isolates were cloned and their nucleotide sequences were determined. The CP similarity (218 amino acids) was highly homologous (>97％) with that of subgroup I CMV strains. However, an additional 20-nulcleotide long segment was only present in 3' NCR of lily isolates, which form an additional stem-loop RNA structure. By using chimeric construct exchange cDNA containing 3'NCR of LK-CMV into the full-length cDNA clone of RNA3 of Fny-CMV, this additional segment may prove to be significant in the identification and fitness of the virus in lily plants. The pathology of zucchini squash infected by F1F2L3-CMV, a pseudorecombinant virus was showed to change drastically the severe mosaic and stunting symptom into a mild chlorotic spot on systemic leave, compared with Fny-CMV. To delimit the sequence of RNA3 affected the pathology, various RNA3 chimeras were constructed between two strains of CMV. The symptom determinants of F1F2L3-CMV were mapped to the positions amino acid 234, 239, and 250 in 3a movement protein (MP). RNA3 chimeras changed the sequences encoding three amino acids were resulted in alteration of systemic symptom.

• 식물병연구
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• 제30권2호
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• pp.157-164
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• 2024

Jack bean (Canavalia ensiformis) is one of healthy products for fermented or functional food in Korea and is widely distributed and cultivated worldwide. During August 2022, Jack bean plants showing symptoms of yellow flecks, chlorosis, necrotic spots and mosaic were observed in Jangheung-gun, South Korea. By transmission electron microscopy, flexuous filamentous virus particles of approximately 750×13 nm in size were observed in the symptomatic leaf samples. The infection of a Korean isolate of clover yellow vein virus (ClYVV-Ce-JH) was confirmed using double antibody sandwich enzyme-linked sorbent assay, reverse transcription polymerase chain reaction and high-throughput sequencing. The complete genome sequence of ClYVV-Ce-JH consists of 9,549 nucleotides (nt) excluding the poly (A) tail and encodes 3,072 amino acids (aa), with an AUG start and UAG stop codon, containing one open reading frame that is typical of a potyvirus polyprotein. The polyprotein of ClYVV-Ce-JH was divided into ten proteins and each protein's cleavage sites were determined. The coat protein (CP) and polyprotein of ClYVV-Ce-JH were compared at the nt and aa levels with those of the previously reported 14 ClYVV isolates. ClYVV-Ce-JH shared 92.62% to 99.63% and 93.39% to 98.05% at the CP and polyprotein homology. To our knowledge, this is the first report of a Korean isolate of ClYVV from Jack bean plants and the complete genome sequence of a ClYVV Jack bean isolate in the world.

• The Plant Pathology Journal
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• 제18권3호
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• pp.121-125
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• 2002

This study was conducted to determine the causal virus that naturally infected hollyhock (Althaea rosea) plant showing mild mosaic symptom in 1999. Flexuous virus particles were found in the cytoplasm of plant tissue from infected hollyhock under transmissible electron microscopy. A virus from the genus Potyvirus under the family Potyviridae was isolated and was maintained on Chenopodium quinoa for three passages. Chlorotic local legions were used to inoculate 20 species of indicator plants. The virus infected all the tested cucurbit plants, but failed to infect Nicotiana benthamiana. Based on the host range test and RT-PCR analysis, the potyvirus was identified as a strain of Zucchini yellow mosaic virus-A (ZYMV-A), one of the major pathogens of cucurbits. Infectivity analysis showed that ZYMV-A induced faster systemic symptom than ZYMV-Cu on squash and other cucurbit plants, suggesting that ZYMV-A was a more severe strain. To better characterize ZYMV-A, Western blot assay was carried rout to the coat protein (CP) of the virus using ZYMV-specific antiserum with ZYMV-Cu and other potyviruses. The CP of the virus reacted strongly with the antiserum against ZYMV, and other tested antisera did not react with the CP of ZYMV-A. Results strongly suggest that the potyvirus infecting hollyhock was a novel strain of ZYMV. This is the first report on ZYMV as the causal virus infecting hollyhock in Korea.

• The Plant Pathology Journal
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• 제35권3호
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• pp.243-256
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• 2019

Field surveys for Plum pox virus (PPV) infection were conducted in stone fruit orchards all over Bulgaria. In total, 1168 out of 3020 leaf samples from cultivated Prunus spp. and wildly growing P. cerasifera trees reacted positive for PPV in DASI-ELISA with the universal monoclonal antibody (MAb) 5B. Further ELISA analyses showed that 987 and 127 isolates belonged to PPV-M and PPV-D serotypes, respectively. The plum and P. cerasifera showed 82.0% and 50.5% levels of infection, respectively followed by the peach (40.0%) and the apricot (32.0%). Five hundred fifty one PPV isolates were further typed by IC-RT-PCR with PPV-Rec, -M and -D-specific primers, targeting (Cter)NIb-(Nter) CP genome region, as 125 isolates were sequenced. The results revealed the presence of PPV-Rec, PPV-M and PPV-D and mixed infections of these strains. PPV-Rec was the most prevalent strain (49.0%), followed by PPV-M (40.1%), while PPV-D was the less spread strain (8.2%). PPV-Rec was the most common strain in plums, including the eight "old-aged" trees from the region of the first Sharka discovery. PPV-M was the most prevalent strain in peach and apricot. Phylogenetic analyses on (Cter)NIb-(Nter)CP of the isolates were performed. PPV-Rec isolates formed a homogeneous group, while PPV-M isolates split into PPV-Ma and PPV-Mb subgroups. Five separated clades were formed by the analyzed PPV-D isolates. Nucleotide sequences of the partial CP coding region of the analyzed isolates revealed a slightly higher intra-strain genetic variability in PPV-Rec and PPV-M isolates, while that of PPV-D strain isolates was higher from the reported for these strains.

• 식물병연구
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• 제9권3호
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• pp.107-115
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• 2003

경상북도 안동, 의성, 군위, 대구의 장마 재배포장에서 모자이크와 괴저반점을 보이는 장마잎을 채집하였다. 이들 이병 잎 조직을 시료로 하여 D N 법 및 ISEM법에 의해 투과전자현미경으로 바이러스 입자를 관찰한 결과, 각각의 시료에서 ChYNMV 항혈청에 반응한 660nm의 사상형 입자가 확인되었다. 장마 잎에서 부분 정제한 바이러스에서 RNA를 추출하여 이것을 주형으로 ChYNMV 특이적 프라이머와 oligo-dT 프라이머를 이용하여 외피단백질 유전자와 3‘-말단 비전사부위를 포함하는 약 1.2kbp의 3’-말단을 증폭하였다. 외피단백질 아미노산서열은 Macluravirus로 알려진 ChYNMV (A B044386)와 97.9%의 상동성을 보여 장마에서 분리한 바이러스를 ChYNMV로 동정하였다. ChYNMV와 다른 Macluavirus의 외피 단백질 아미노산서열을 비교한 결과, M말단 영역에서 가장 많은 변이를 확인하였고 Macluravirus 속에는 잘 보존된 영역이 존재하였다.

• 식물병연구
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• 제24권4호
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• pp.285-291
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• 2018

국내육성딸기 품종인 설향과 감홍에서 딸기누른오갈바이러스의 국내 분리주 2종을 분리하고 외피단백질 전체 염기서열을 결정하고 분석하였다. 국내 분리주 SH와 KH의 외피단백질 염기와 아미노산 상동성은 각각 90.4%와 95.5% 였다. 기존에 국내에서 보고된 KNS1분리주와 GenBank에 등록된 45개의 다른나라 분리주 외피단백질 염기서열을 모두 수집하여 총 48개 SMYEV 외피단백질에 대한 계통학적 유연관계를 분석할 결과 총 5개의 subgroup (I-V)으로 분류가 되었다. 이 중 subgroup IV과 V과 새로운 변이집단으로 국내분리주도 KH와 KNS1은 subgroup I에 포함된 반면, SH는 새로운 subgroup인 IV에 포함되어 국내분리주간에도 계통이 다른 것을 추측할 수 있었다. 유전적 다양성 분석결과 SMYEV의 새로운 subgroup의 다양성이 더욱 높은 것으로 나타나 SMYEV가 유전적으로 진화를 하고 있음을 알 수 있었다. 이 논문은 국내 SMYEV 분리주에 대한 분자적 특성에 대한 첫 보고이다.

• 식물병연구
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• 제30권1호
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• pp.60-65
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• 2024

2020년 7월, 전라남도 해남에서 퇴록병반과 엽맥녹대 등 바이러스 병징이 보이는 패션프루트(Passiflora edulis) 잎으로부터 total RNA를 추출했으며, RT-PCR 검정과 염기서열 결정을 통해 CMV-HN2를 동정하였다. 패션프루트에 감염하는 CMV의 생물학적 특성을 확인하기 위해 10개 지표식물에 CMV-HN2를 접종한 결과, CMV-Fny와 비교하여 전형적인 CMV의 감염 패턴을 나타냈다. CMV 패션프루트 분리주들의 외피단백질에 대한 계통발생학적 분석한 결과, CMV의 패션프루트 분리주들은 subgroup I에 속하는 것으로 나타났으며, 그중 CMV-HN2는 subgroup IA에 속하는 것으로 나타났다. 또한 국내 패션프루트 CMV 분리주들 사이의 subgroup 차이가 확인되었다. 패션프루트 분리주 간의 외피단백질 아미노산 서열을 비교하였을때, CMV-HN2는 다른 분리주들과 특이적으로 4-8개 아미노산 차이가 존재하였다. 이 결과를 통해, 패션프루트 CMV 분리주들의 외피단백질에서 유전적 다양성이 있음을 확인하였다.

• The Plant Pathology Journal
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• 제33권4호
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• pp.393-401
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• 2017

Efforts to control viral diseases in crop production include several types of physical or chemical treatments; antiviral extracts of a number of plants have also been examined to inhibit plant viral infection. However, treatments utilizing naturally selected microorganisms with activity against plant viruses are poorly documented. Here we report isolation of a soil inhabiting bacterium, Pseudomonas oleovorans strain KBPF-004 (developmental code KNF2016) which showed antiviral activity against mechanical transmission of tobamoviruses. Antiviral activity was also evaluated in seed transmission of two tobamoviruses, Pepper mild mottle virus (PMMoV) and Cucumber green mottle mosaic virus (CGMMV), by treatment of seed collected from infected pepper and watermelon, respectively. Pepper and watermelon seeds were treated with culture supernatant of P. oleovorans strain KBPF-004 or control strain ATCC 8062 before planting. Seeds germinated after treatment with water or ATCC 8062 yielded about 60% CGMMV or PMMoV positive plants, whereas < 20% of KBPF-004-treated seeds were virus-infected, a significantly reduced seed transmission rate. Furthermore, supernatant of P. oleovorans strain KBPF-004 remodeled aggregation of PMMoV 126 kDa protein and subcellular localization of movement protein in Nicotiana benthamiana, diminishing aggregation of the 126 kDa protein and essentially abolishing association of the movement protein with the microtubule network. In leaves agroinfiltrated with constructs expressing the coat protein (CP) of either PMMoV or CGMMV, less full-size CP was detected in the presence of supernatant of P. oleovorans strain KBPF-004. These changes may contribute to the antiviral effects of P. oleovorans strain KBPF-004.