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題名 | 牛傳染性鼻氣管炎病毒(臺灣YL株)免疫原gI基因之分子選殖=Molecular Cloning of the Immunogen gI Gene of Infectious Bovine Rhinotracheitis Virus Strain YL |
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作者姓名(中文) | 鍾楊聰; 徐維莉; | 書刊名 | 臺灣畜牧獸醫學會會報 |
卷期 | 66:1 1996.03[民85.03] |
頁次 | 頁7-17 |
分類號 | 437.3 |
關鍵詞 | 牛傳染性鼻氣管炎病毒; 封套醣蛋白gI; 基因選殖; Infectious bovine rhinotracheitis virus; Envelope glycoprotein gI; Gene cloning; |
語文 | 中文(Chinese) |
中文摘要 | 本篇報告完成台灣分離YL株牛傳染性鼻氣管炎病毒(infectious bovine rhinotracheitis virus)封套醣蛋白(envelope glycoprotein)gI引基因之選殖。我們使用 限制酵素(restriction enzyme)HindⅢ和BamHI併同切割病毒基因體核酸(genomic DNA), 經0.8%低熔點瓊脂電泳(low-melting point agarose gel electrophoresis)切下大小在 8-12kb左右的所有DNA片段,並將DNA溶出,再以黏合酵素(T4 DNA ligase)和已經相同限 制酵素切割過的pBR322選殖載體DNA相黏合。按著將DNA樣本送入大腸桿菌勝任細胞 (competent cells) HB101中使轉形,初步篩選結果得到五個嵌有不同病毒核酸片段的重組 質體(recombinant plasmid)。隨後我們根據國外已發表的美國Cooper株病毒刨gI基因序列 設計並合成了一對寡核甘酸引子(oilgonucleotide primers),用以進行聚合酵素連鎖反應 (Polymerase chain reaction)從五個質體中篩選出含有gI基因的質體。經進一步限制酵素 切割分析與核甘酸定序分析(nucleotide sequence analysis),確認我們所選殖到的質體具 有11.7kb嵌入片段(insert);長的3kb的完整gI基因居於中段位置,兩側翼序列(flanking sequence)各有5.2和3.5kb長。如此長度符合同源型互換(homologous recombination)發 生的要件,足供將來建構基因缺損病毒株(genedeleted live virus)之用。 |
英文摘要 | Infectious bovine rhinotracheitis virus (IBRV) is a causative agent of infectious bovine rhinotracheitis in cattle. The virus synthesizes at least seven envelop glycoproteins and four of them have been well characterized and named go (gB), gII (gE), gIII (gC), and gIV (gD). These four major glycoproteins are the major target antigens involved in virus neutralization. Previously, we have cloned and completed the nucleotide sequence analysis of the genes encoding the glycoproteins gIII and gIV of IBRV strain YL isolated in Taiwan. In this report, we present the molecular cloning of the IBRV gI gene. A 11.7 kilobase-pair (kb) HindIII-BamHI fragment derived from viral genome was cloned into a pBR322 plasmid vector. The result of analysis by the polymerase chain reaction, of which the primers were designed according to the published gI gene sequence. Subsequent restriction mapping, subcloning, and partial nucleotide sequence analysis on the 11.7-kb fragment further verified the identity of the IBRV gI gene. |
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