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題 名 | 胡瓜嵌紋病毒感染引起的港口馬兜鈴嵌紋病之鑑定=Identification for the Infection of Cucumber Mosaic Cucumovirus on Aristolochia Zolligeriana |
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作 者 | 陳金枝; 張清安; 林玫珠; 方懷盛; | 書刊名 | 植物病理學會刊 |
卷 期 | 9:1 2000.03[民89.03] |
頁 次 | 頁29-34 |
分類號 | 435.13 |
關鍵詞 | 港口馬兜鈴; 胡瓜嵌紋病毒; 鑑定; PCR偵測; 核酸探針; 雜配反應; Aristolochia zolligeriana; CMV; Identification; PCR detection; DNA probe; Hybridization; |
語 文 | 中文(Chinese) |
中文摘要 | 胡瓜嵌紋病毒感染引起的港口馬兜鈴嵌紋病之鑑定 |
英文摘要 | The aristolochia zolligeriana Miq. (AZO), a native species of Aristolochiaceae in Taiwan, is the major victual plant of Papilio spp. butterfly in nature. During artificial cultivation of AZO, some plants exhibiting systemic mosaic symptoms were found in one of the experiment plantation in Taiwan Endemic species Research Institute. A virus isolate (Az1) was obtained from the diseased AZO plants by three successive local-lesion selections on Chenopodium quinoa and subsequently propagated in Nicotiana benthamiana. The same mosaic symptom as that on the original diseased plants was reproduced on healthy AZO seedlings by inoculation with Az1, indicating the Az1 should be the causal agent of AZO mosaic disease. Besides AZO, Az1 could infect N. benthamiana and N. tobacum cv. Xanthi and induce systemic mosaic symptoms. By electron microscopy, isometric particles with approximately 30 nm in diameter was observed in negatively stained sample obtained from clarified viral concentrate (CVC) procedure described by Christie et al. (1987). In ELISA test, field collected tissue from diseased AZO, Az1-inoculated tissues of N. benthamiana and AZO were all strongly reacted with the antiserum against cucumber mosaic cucumovirus (CMV) but not with antiserum against bean yellow mosaic potyvirus (BYMV) and Potyvirus-specific monoclonal antibody (Agdia Inc.). In addition, these antigens were all indistinguishable to the control CMV agtigen when tested in sodium dodecyl sulfate-immunodiffusion test. Using a primer pair designed for specific amplification of CMV, a 486-bp DNA fragment equivalent to the one amplified from the control CMV, was produced from Az1-infected tissues by reverse transcription polymerase chain reaction (RT-PCR). The same primer pair could also amplify tomato aspermy cucumovirus (TAV), a virus serologically related to CMV, but produced a DNA fragment larger than that amplified from CMV. However, only the DNA product amplified from Az1-infected tissue, but not the one amplified from TAV, could be hybridized with the biotin-labeled DNA probe of CMV. Furthermore, a primer pair for specific amplification of TAV could not amplified any product from Az1-infected tissues, confirming that Az1 was an isolate of CMV instead of TAV. When the 486-bp DNA fragment was cloned and sequenced, it was found corresponding to 3'-terminal region of coat protein gene and part of the 3'-nontranslated region of CMV RNA 3. Comparing with the known CMV sequences from the Genebank, this Az1 DNA fragment shared 92-93% and 98% of identities in the nucleotide and deduced amiono acid sequence, respectively. Based on the biological, serological and molecular properties of Az1 as revealed in this study, the virus should reasonably be recognize as an isolate of CMV. To our knowledge, this is the first report of A. zolligeriana as a natural host species for CMV. |
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