Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/31035
標題: 洋桔梗壞疽病毒彩色海芋分離株之鑑定與特性分析
Identification and characterization of a calla lily-infecting isolate of Lisianthus necrosis virus
作者: 張怡珊
Chang, Yi-Shan
關鍵字: Lisianthus necrosis virus
洋桔梗壞疽病毒
Zantedeschia spp.
彩色海芋
出版社: 植物病理學系所
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摘要: 台灣中部地區田間栽培的彩色海芋 (calla lily, Zantedeschia spp.),發現疑似由病毒感染所引發的系統性壞疽病徵之植株。田間彩色海芋病株呈系統性感染,葉緣捲起,葉面出現黃化斑及壞疽斑,未壞死處略呈嵌紋狀,嚴重者全葉壞疽或整株死亡。罹病葉片粗汁液、純化的懸浮液中及病組織超薄切片,經以電子顯微鏡觀察到一直徑約為 32-34 nm的球形病毒顆粒。間接酵素聯結免疫吸附分析法 (indirect-ELISA)、西方轉漬免疫分析 (western blotting) 試驗均顯示受測的彩色海芋病毒與洋桔梗壞疽病毒 (Lisianthus necrosis virus, LNV) 有血清類緣關係。電泳分析 (SDS-PAGE) 彩色海芋分離株得到一分子量約 39 kDa 的鞘蛋白,此與 LNV 的鞘蛋白分子量相似。進一步針對洋桔梗壞疽病毒彩色海芋分離株 (LNV-Z) 的基因體進行選殖及核酸序列之解析。將LNV-Z 繁殖於其系統性寄主 Nicotiana benthamiana 上,經抽取 LNV-Z 總量 RNA 後,針對其基因體序列,以反轉錄聚合酶鏈鎖反應 (RT-PCR) 進行增幅並選殖所增幅的 cDNA 片段,經解序得知 LNV-Z 基因體全長度為 4785 個核苷酸 (Accession number AM71119)。基因體由 5 個轉譯架構 (ORFs) 所組成,由 5′ 端至 3′ 端分別為 ORF 1 (聚合酶基因,polymerase gene, 2457 nt)、ORF 2 (外鞘蛋白基因,coat protein gene, 1167 nt)、ORF 3 (移動蛋白基因,movement protein gene, 576 nt)、ORF 4 (p19蛋白基因,p19 protein gene, 519 nt)。此基因體結構與 LNV 和其他Tombusvirus 屬病毒相似。序列分析的結果顯示: LNV-Z與 Tombusvirus 屬病毒的鞘蛋白胺基酸相同度 (amino acid identity) 為 36.1~97.7%,其中以與 LNV 洋桔梗分離株 (LNV-L) 的相同度最高(鞘蛋白胺基酸序列相同度 97.7%)。由此分子層次證據進一步顯示引起彩色海芋系統性壞疽病徵的病原確定為洋桔梗壞疽病毒 (LNV)。
Calla lily (Zantedeschia spp.) showing symptoms of systemic necrosis was observed in the fields of central Taiwan in the 2003. Electron microscopic observation revealed the presence of isometric virus particles of 32-34 nm in diameter in the preparations of crude extracts, purified virus suspension and ultrathin sections of diseased tissues. The relative molecular weight of the subunit of viral capsid protein, similar to that of Lisianthus necrosis virus (LNV), was estimated as 39 kDa by SDS-polyacrylamide gel electrophoresis. The isolated virus reacted positively to LNV antiserum in indirect-ELISA and western blotting tests. The cDNA fragments were amplified from the total RNA extracted from LNV-Z infected Nicotiana benthamiana by reverse transcription-polymerase chain reaction (RT-PCR). Full length sequence of the virus genome has been compiled from cDNA fragments amplified by RT-PCR with LNV-specific primer sets. Sequence analysis showed that the complete genome of LNV-Z consists of 4785 nucleotides with five open reading frames (Acc. No. AM71119). The amino acid identity of the coat proteins between tested strain (LNV-Z) and the reported lisianthus strain (LNV-L) was 97.7%. The results shown in this study indicate that LNV is the causal agent of the systemic necrosis of calla lily in the fields.
URI: http://hdl.handle.net/11455/31035
其他識別: U0005-2208200716274600
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