Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/31423
標題: 臺灣不同地區木瓜輪點病毒西瓜型分離株鞘蛋白基因之分析
Analyses of the Coat Protein Genes of Papaya Ring Spot Virus W Type Isolates from Different Areas of Taiwan
作者: 王如蓉
Wang, Ju-Jung
關鍵字: Papaya ring spot virus W
木瓜輪點病毒
PRSV W
coat protein
鞘蛋白
出版社: 植物病理學系
摘要: 木瓜輪點病毒 (papaya ringspot virus, PRSV) 屬於馬鈴薯 Y 病毒 屬 (The Potyvirus genus), 其寄主範圍可分為西瓜型 (PRSV W) 和木瓜 型 (PRSV P) 二型。西瓜型 (W) 可感 染葫蘆科和藜科植物,在許多葫蘆 科之瓜類作物危害嚴重。而木瓜型 (P) 除了可感染葫蘆 科及藜科外,尚 能再感染番木瓜科,造成木瓜的嚴重危害。 為了比較台灣不同地區木瓜 輪 點病毒西瓜型之鞘蛋白基因變異情形, 乃自各地收集六種瓜類作物共 231 個樣本, 其中 12 個分離株經由血清及寄主反應之測試,證實確為 PRSV W 型分離株,其樣本分別來自台 中,彰化,雲林,嘉義,台南及屏 東。這 12 個 W 分離株之鞘蛋白基因經由反轉錄聚合脢 連鎖反應 (RT- PCR) 放大之後,以三種核酸限制酵素 RsaI, AluI 和 NIaIII 剪切後, 比 較其電泳圖譜,發現其圖譜變化性不大, 且和 P 型 YK 分離株 ( 分 自台南 ) 及 P-PD ( 泰國 ) 分離株之圖譜相似。 從此多形性圖譜來看 , 台灣之 PRSV W 分離株和來自台灣之 PRSV P-YK 與泰國之 P-PD 較相 近。 進一步以反轉錄聚合脢連鎖反應選殖嘉義、台南、屏 東三地 W 型 分離株 (W-CI, W-TN, W-PT) 鞘蛋白基因的選殖株,解出其核甘酸序列, 並比 較其異同。 三者之鞘蛋白基因之核甘酸相同度為 96.37-96.99 %, 氨基酸相同度為 94.46-96.74 %, 而在 3' 端非轉譯區 (3 ? non-coding region) 之核甘酸相同度達 98.56-99.04 %。這顯示 W-CI ,W-TN 和 W-PT 三個分離株之關係相近,且變異性不大。而 再比較這三 個 W 型分離株和其他已發表之 W 型 FL ( 佛羅里達 ), W-AU ( 澳洲 ), P 型 P-YK ( 台南永康 ),P-HA ( 夏威夷 ),P-FL ( 佛羅里達 ) 在鞘蛋白基因之核甘酸及 氨基酸相同度上作比較, 則發現 W-CI, W- TN 和 W- PT 和 P-YK 在核甘酸上有 95.93-96.64 % 的相同度, 在氨基酸上有 95.21- 96.16 % 的相同度, 但和其他不同來源 ( 非台灣 ) 之 W 和 P 型比較,核甘酸相同度只有 90.59-91.97 %。 而非台灣之 W 和 P 型分離株之間,W-AU,W-FL, P-HA,P-FL 之氨基酸相同度則為 96.12-99.3 %。在 3' 端 非轉譯區之核甘酸比較上, W-CI,W-TN, W- YK 三個台灣 W 型分離株和來自台灣的 P-YK 有 97.21-98.08 % 的相同 度, 而和不來自台灣的 W 和 P 型分離株則只有 88.51-93.3 % 的相同 度。這些結果顯示 W-CI,W-TN 和 W-PT 三個台灣的 W 型彼此間在鞘蛋 白基因變異 情形不大, 且和同樣來自台灣的 P-YK 較相近而和來自美、 澳系統的 W 和 P 型關係較遠 , 本研究之結果顯示其類緣關係之演化乃 視病毒系統地域性之差異而非依其病原性來決定 。
Papaya ringspot virus (PRSV), a member of the Potyvirus genus, contains two major groups, type P ( PRSV P) and type W (PRSV W) virus. The host range of PRSV W strains is limited to Chenopodiaceae and Cucurbitaceae, whereas P type strains infect Cariceae (papaya) in addition. In order to develop transgenic cucurbits with resistance to PRSV W in Taiwan, typical PRSV W isolates from the island were collected and the variability in the nucleotide and amino acid sequences of the CP genes were analyzed. Twelve isolates from the forty ELISA positive samples, collected from six cucurbit crops from different areas of Taiwan, were identified by host reactions and serology tests as typical W type virus isolates. They did not infect papaya but were serologically indistinguishable from type P isolates when tested against the antisera to PRSV P type or PRSV W type. In order to analyze the polymorphisms of the CP genes of the 12 PRSV W type isolates from Taiwan and other reported PRSV W and P type viruses, digestion patterns of RT-PCR products,which were amplified from the N-terminal half of the CP genes, with RsaI, AluI and NlaIII were compared. The results showed that most of the 12 PRSV W isolates from Taiwan were closely related to PRSV P- YK, a P type strain from Taiwan, and P-PD, a P type strain from Thailand,and far apart from P-HA, a P type strain from Hawaii, and W-FL, a W type strain from Florida. To further analyze the variation of the CP genes of PRSV W isolates from Taiwan, three isolates, PRSV W-CI (from Chiayi), W-TN (from Tainan) and W-PT ( from Pingtung), were chosen for cDNA cloning for determination of their CP gene sequences. Comparison the three isolates of Taiwan with each other, indicated that they shared 96.37-96.99% and 94.46-96.74% nucleotide and amino acid identity, respectively, and the nucleotide identity of the 3' non-coding regions were 98.56-99.04%, indicating that the three isolates were closely related strains of the same virus. Comparison of the three W isolates from Taiwan with other reported W isolates of W-FL (from Florida) and W-AU (from Australia), and P isolates of P- YK (from Yung Kang, Taiwan), P-HA (from Hawaii), and P-FL (from Florida) revealed that the nucleotide identity of the CP genes of the three W type isolates from Taiwan shared higher percentages of 95.93-96.64% with P-YK, another P type isolate from Taiwan, they shared lower percentages of 90.59-91.97% with other non-Taiwan PRSV W and P type isolates. In terms of amino acid identity of the CP genes, the three PRSV W type isolates from Taiwan, shared 95.21-96.16% with PRSV P- YK. The comparison of non-Taiwan W and P type isolates showed that they had 95.12-99.3% amino acid identity. The comparison of the 3' non-coding regions of the eight PRSV isolates, revealed that the three PRSV W isolates from Taiwan shared 97.21-98.08% nucleotide identity with P-YK, whereas they shared only 88.51-93.3% nucleotide identity with other non-Taiwan PRSV P and W isolates. The sequences analyses indicated that the relationships among these W and P type isolates could be divided into two groups, one contained PRSV W-CI, W-TN, W-PT and P-YK, the other one contained PRSV W-AU, W-FL, P-HA and P-FL. These coupled with enzyme-digestion polymorphisms indicated that the CP genes of the three W type isolates from Taiwan were closely related to P-YK(a Taiwan isolate) and far apart from W and P type isolates in other different geographic areas. Our results also implicate that degrees of variation of the CP genes do not follow major differences in host specificity, such as papaya or non-papaya infecting, but are more closely corresponding to geographic distribution.
URI: http://hdl.handle.net/11455/31423
Appears in Collections:植物病理學系

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