• The Plant Pathology Journal
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• 제21권2호
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• pp.167-171
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• 2005

We isolated the Cucumber green mottle mosaic virus(CGMMV) particles from watermelon leaves and designated as CGMMV-HY1 as a watermelon isolate and attempted to characterize the pathogenic isolate responsible for such an epidemic in watermelon and also to monitor dominant viral isolates in greenhouse. The watermelon plants infected with CGMMV generally showed mottle mosaic, mosaic, growth stunting, necrosis and deformed fruit. The reactions of indicator plants to CGMMV-HY1 were the local lesions on Nicotiana tabacum cv. White Burley, Nicotiana tabacum cv. Samsun, and Chenopodium amaranticola, and the mosaic symptoms only on Cucumis sativus, but the CGMMV-HY1 did not infect Nicotiana sylvesytis, Datura stramonium, Chenopodium quinoa, and Petunia hybrida. Purified virus particles were rod-shaped and about 300 nm long. The coat protein (CP) of purified CGMMV-HY1 was single band with molecular weight of about 16.5 kDa which was confirmed by western blot analysis probed with monoclonal antibody of CGMMV-HY1. The genomic and subgenomic RNAs of 6.4 kb and 0.75 kb were revealed by the electrophoresis on 1.2% formaldehydedenatured agarose gel. Viral and complementary CGMMV-specific primer sets were designed for spanning the genome using previously reported CGMMV sequences. A 464bp of CP gene of CGMMV-HY1 was amplified by RT-PCR and cloned into PGEM-T easy vector. The nucleotide sequence of CP gene of CGMMV-HY1 shared 98%, 99%, and 100% identities with that of CGMMV strains W, KOM, and KW respectively. Based on these results, we identified CGMMV-HY1 as a CGMMV isolate of watermelon, a member of Tobamovirus.

• Journal of Microbiology and Biotechnology
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• 제26권1호
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• pp.160-170
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• 2016

The increasing spread of antibiotic-resistant pathogens has raised the interest in alternative antimicrobial treatments. In our study, the functionally active gram-negative bacterium bacteriophage CP933 endolysin was produced in Nicotiana benthamiana plants by a combination of transient expression and vacuole targeting strategies, and its antimicrobial activity was investigated. Expression of the cp933 gene in E. coli led to growth inhibition and lysis of the host cells or production of trace amounts of CP933. Cytoplasmic expression of the cp933 gene in plants using Potato virus X-based transient expression vectors (pP2C2S and pGR107) resulted in death of the apical portion of experimental plants. To protect plants against the toxic effects of the CP933 protein, the cp933 coding region was fused at its Nterminus to an N-terminal signal peptide from the potato proteinase inhibitor I to direct CP933 to the delta-type vacuoles. Plants producing the CP933 fusion protein did not exhibit the severe toxic effects seen with the unfused protein and the level of expression was 0.16 mg/g of plant tissue. Antimicrobial assays revealed that, in contrast to gram-negative bacterium E. coli (BL21(DE3)), the gram-positive plant pathogenic bacterium Clavibacter michiganensis was more susceptible to the plant-produced CP933, showing 18% growth inhibition. The results of our experiments demonstrate that the combination of transient expression and protein targeting to the delta vacuoles is a promising approach to produce functionally active proteins that exhibit toxicity when expressed in plant cells.

• Journal of Plant Biology
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• 제34권2호
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• pp.101-106
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• 1991

담배(N. glauca) 잎 절편 유래 callus 배양에 있어서 callus 생장 단계에 따라 nicotine 생성에 미치는 두 가지 auxin (2,4-D와 NAA)의 영향을 HPLC를 이용하여 조사하였다. 고농도($11.5\;\mu\textrm{M}$)의 2,4-D는 배양 4주째에, NAA는 배양 2주째에 nicotine 생성을 촉진시켜 최대함량을 나타내었고, 그 이후로는 감소하였다. 새성된 nicotine이 다른 alkaloid로 전환되었다. 한편, 전구물질로 알려진 L-arginine 또는 L-aspartic acid 첨가배양에서 저농도($1.5\;\mu\textrm{M}$)의 2,4-D는 고농도에 비해 nicotine 생성을 촉진시켰고, 저농도의 NAA는 고농도에 비해 억제시켰다. 그러나 L-aspartic acid 첨가배양에서 저농도의 NAA는 배양 5주까지 nicotine 생성을 촉진시켰다. 이와 같은 결과로부터 두 가지 auxin은 nicotine 생성에 대해 효과가 다르며, L-aspartic acid가 nicotine 생합성의 전구물질로서, 저농도의 NAA는 이 합성경로에 작용하여 nicotine 생성을 유도하였음을 추론할 수 있었다.

• Molecules and Cells
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• 제27권1호
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• pp.47-54
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• 2009

A cDNA clone for a transcript preferentially expressed during an early phase of flooding was isolated from Nicotiana tabacum. Nucleotide sequencing of the cDNA clone identified an open reading frame that has high homology to the previously reported glycine-rich RNA-binding proteins. The open reading frame consists of 157 amino acids with an N-terminal RNA-recognition motif and a C-terminal glycine-rich domain, and thus the cDNA clone was designated as Nicotiana tabaccum glycine-rich RNA-binding protein-1 (NtGRP1). Expression of NtGRP1 was upregulated under flooding stress and also increased, but at much lower levels, under conditions of cold, drought, heat, high salt content, and abscisic acid treatment. RNA homopolymer-binding assay showed that NtGRP1 binds to all the RNA homopolymers tested with a higher affinity to poly r(G) and poly r(A) than to poly r(U) and poly r(C). Nucleic acid-binding assays showed that NtGRP1 binds to ssDNA, dsDNA, and mRNA. NtGRP1 suppressed expression of the fire luciferase gene in vitro, and the suppression of luciferase gene expression could be rescued by addition of oligonucleotides. Collectively, the data suggest NtGRP1 as a negative modulator of gene expression by binding to DNA or RNA in bulk that could be advantageous for plants in a stress condition like flooding.

• 미생물학회지
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• 제1권1호
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• pp.10-18
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• 1963

Studies with the Tobacco Mosaic Viruses; W. S Kim, and So, I Y., (Dept. of biology Sung Kyun Kwan Univer. Seoul, Korea.). Using the common strain of tobacco mosaic virus (TMV) which was sent from the Dept. of Plant Pathology, University of Wisconsin, U.S.A. as control virus, a possible new strain of tobacco mosaic virus (SMV) was isolated from tobacco leaves collected from Tobacco Experiment Station farms as well as from various blends of manufactured Korean cigaretts. SMV was isolated by single lesion isolation method and by inoculating the virus through various species of host plants. The two viruses, TMV and SMV were indentified by the difference in symptoms, host range, serological reaction, and electron micrograpy. As the results of the above experiment the author believes the virus isolate SMV is a different strain of TMV. The experimental evidences that SMV belongs to the TMV group are as follows; 1. Both viruses produced local necrotic lesions on Nicotiana glutimosa L. 2. Both showed a dilution end point of $10^8$. 3. Aphid transmission was failed with the viruses. 4. Both had an isoelectric point around pH 3.3. 5. Two viruses were serological reactive. 6. The size of the virus particles was around 270-300mu as they were observed under the electron microscope. The virus SMV, however, is different from the common strain of TMV and the experimental evidences are as follows; 1. SMV produced quite different symptoms from TMV on various host plants like tobacoo(Nicotiana tabacum L., White Burley), Nicotiana rustica L., Chenopodium Koreanse Nakai. Bata vulgaris L., and Datura tatula L., SMV produced distinct local lesions on these host plants whereas TMV incited largely mosaic diseases. 2. The serological titers obtained from the heterologous combinations were lower than those from homologous combinations of antigens and antiser.

• The Plant Pathology Journal
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• 제22권4호
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• pp.353-359
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• 2006

Transgenic Nicotiana benthamiana plants harboring the coat protein(CP) gene of Zucchini green mottle mosaic virus(ZGMMV) were chosen as a model host for the environmental risk assessment of genetically modified plants with virus resistance. This study was focused on whether new virus type may arise during serial inoculation of one point CP mutant of ZGMMV on the transgenic plants. In vitro transcripts derived from the non-functional CP mutant were inoculated onto the virus-tolerant and -susceptible transgenic N. benthamiana plants. Any notable viral symptoms that could arise on the inoculated transgenic host plants were not detected, even though the inoculation experiment was repeated a total of ten times. This result suggests that potential risk associated with the CP-expressiing transgenic plants may not be significant. However, cautions must be taken as it does not guarantee environmental safety of these CP-mediated virus-resistant plants, considering the limited number of the transgenic plants tested in this study. Further study at a larger scale is needed to evaluate the environmental risk that might be associated with the CP-mediated virus resistant plant.

• KSBB Journal
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• 제22권5호
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• pp.313-317
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• 2007

본 연구에서는 Pluronic F-68을 해동 후 회복배지에 적용하여, 동결보존된 형질전환 식물세포의 생장 증진을 도모하였다. Pluronic F-68은 세포표면의 소수성을 감소시켰을 뿐만 아니라, 세포의 투과성을 증진시켜 세포와 직접 상호 작용할 수 있음을 관찰하였다. 또한 Pluronic F-68의 첨가에 의해 재조합단백질의 발현에 부정적인 영향을 보이지 않음을 확인하였다. 따라서 동결보존시 해동 후 회복배지에 Pluronic F-68의 첨가는 식물세포의 신속하고 효율적인 대량 현탁배양을 가능하게 할 수 있다.

• 한국응용곤충학회지
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• 제20권4호
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• pp.191-195
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• 1981

모자익병징을 나타내는 호박, 박, 오이, 참외, 수박의 박과작물에서 Watermelon Mosaic Virus(WMV)를 분리동정하였다. 이병주로 부터 분리된 WMV는 복숭아혹진딧물(Myzus persicae, Sulzer)에 의해 쉽게 충매전염되었고 종자전염은 되지 않았다. 지표식물검정결과 명아주에서는 접종엽에 대형의 국부병반이, 호박, 박, 오이, 참외, 수박에서는 접종상엽에서 모자익병반이 나타났고 담배(Nicotiana tabacum, Bright yellow), 담배 (Nicotanan glutinosa), 동부, 페추니아, 독말풀에서는 반응이 나타나지 않았다. WMV isolate의 물리적 성질은 내열성 $55\~65^{\circ}C$, 내희석성 $10^{-4}\~10^{-5}$배, 내보존성 $7\~8$일간이었다. WMV isolate의 항혈청반응은 미량침강법에서 양성으로 나타났고 바이러스립자의 형태는 전자현미경검경결과 대부분이 750nm의 사상입자이었다. 이상의 시험결과로 박과 작물에서 분리한 바이러스가 WMV 임이 확인되었다.

• 식물조직배양학회지
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• 제21권6호
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• pp.333-339
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• 1994

연초의 약배양에 의해서 반수체를 유기하고 형성된 반수체와 이배체의 캘러스로부터 식물체 분화과정에서 일어나는 생리적 차이점을 비교하고자 효소의 활성도와 페놀화합물의 함량을 비교 조사하였다. 반수체의 형성은 화아가 중간정도된 크기를 이용하여 IAA가 1.0 mg/L kietin이 0.5mg/L 조합 처리되고 활성탄이 3 mg/L 함유된 배지에서 가장 양호하였다. 캘러스의 유도는 이배체 및 반수체 공히 2,4-D의 농도가 0.5 mg/L 첨가된 배지에서 양호하였으며, 캘러스로부터 재분화는 BAP 농도 2.0 mg/L에서 효과적이었다. 재분화시 변화되는 효소의 활성도와 페놀화합물의 함량은 처리된 BAP의 농도와 사용한 식물체에 따라 차이가 있었다. Peroxidase의 활성도는 이배체 및 반수체 공히 BAP 농도가 2.0 mg/L일때 가장 높았으며, catalase의 활성도는 BAP의 농도가 1 mg/L일때 가장 높은 경향을 보였다. IAA oxidase 및 catalase의 활성도는 이배체에 비해 반수체에서 더 높은 경향을 보였으나 peroxidase의 활성도는 이배체가 더 높은 경향을 나타내었다.

• Journal of Plant Biology
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• 제38권2호
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• pp.203-209
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• 1995

담배(Nicotiana tobacum L. cv. Wisconsine 38) 잎 절편을 해녀콩(Canavalia lineata)의 leghemoglobin(Lb) cDNA를 포함하는 Agrobacterium과 함께 배양한 후, 0.5mg/L BAP, 0.1mg/L ${\alpha}-NAA$와 200mg/L kanamycin, 500 mg/L carbenicillin을 포함하는 MS 배지에서 선별하여 7개의 재분화 개체를 얻었다. 이로부터 분리한 게놈 DNA에 대한 Southern 혼성화 반응과 PCR 결과로 Lb cDNA가 담배의 게놈에 삽입되었음을 확인하였다. 형질전환된 담배로부터 분리한 RNA에 대한 northern 혼성환 실험 결과 약 1,000 nt의 RNA가 혼성화 반응을 보였으며, 총 RNA를 주형으로 합성한 1차 가닥의 cDNA를 PCR로 증폭한 결과, Lb cDNA와 혼성화 반응을 보이는 0.5 kb의 DNA가 증폭되었다. 콩의 Lb에 대한 다군항체(polyclonal antibody)를 사용하여 단백질 면역 항체 반응을 실시한 결과, Lb로 판단되는 약 15.8 kD의 위치에서 혼성화 반응이 나타났다. 이상의 결과로 해녀콩 Lb cDNA가 형질전환된 담배의 게놈에 삽입되었을 뿐만 아니라 mRNA로 전사되어 Lb 단백질로 해독되었음을 알 수 있었다.