Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/21654
標題: 立枯絲核菌線形質體之限制�}圖譜分析與選殖定序
Mapping and sequencing of linear plasmid DNA in Rhizoctonia solani
作者: 曾雅秀
Tseng, Yai-hsiu
關鍵字: Rhizoctonia solani
立枯絲核菌
linear plasmid
restriction map
線形質體
限制圖譜
出版社: 植物學系
摘要: 本研究以本省的立枯絲核菌(Rhizoctonia solani)第四融和群的強致病性 CHU352為材料, 對其線形、分子量約 2.6 kb 質體 pCHU352進行限制圖 譜的分析與DNA定序。此質體位於於粒線體中, 利用CTAB法可將質體DNA與 染色體DNA一起由細胞中抽出,而利用洋菜膠電泳將之分離; 經過限制( PvuⅡ, BclⅠ, KpnⅠ, EcoRⅠ, SmaⅠ, BglⅡ, BamHⅠ )切割後, 此 2.6 Kb 大小的質體被略分為三種型式。此三種型式的質體DNA除了EcoRⅠ 切位外, 並不具有其他共同切位。 pCHU352在 proteinase K 處理後, 對 exonucleaseⅢ有不敏感性(insensitive), 顯示其端點並非結合蛋 白( binding protein); 經變性處理後分析其pattern大小及位置並無變 化,且質體對S1 nuclease不敏感, 間接證明了兩端的髮夾形結構。 pCHU352經AvaⅠ, HindⅢ, SacⅠ, XhoⅠ切割後分別被選殖在 E.coli 的 pBluescript KS(+) 或 pGEM-3zf(+),其中 1.5Kb大小的 HindⅢ片段 已被定序並與另一強制病性菌株CHU341的質體做比較。
Plasmid pCHU352 is a linear DNA element of Rhizoctonia solani AG4, a strong pathogenic strain, CHU352. The molecular weight of pCHU352 is 2.6 Kb. This study was tried to analyze the restriction maps of plasmid pCHU352 and clone the restriction fragments from pCHU352 for DNA sequencing. Restriction map of the plasmid was constructed by digested with different enzymes ( PvuⅡ, BclⅠ, BamHⅠ, KpnⅠ, EcoRⅠ, BglⅡ, SmaⅠ), and the result suggested that there exist three types of plasmids. Distribution of the restriction sites were different among the three plasmid DNAs except the EcoRⅠ cutting site. The plasmid DNAs were resisted to exonuclease Ⅲ after treated with proteinase K, in contrast, treated with S nuclease will make the plasmid DNAs be sensitive to exonuclease Ⅲ. This result indicated that the linear plasmid has hairpin loops, not terminial binding protein at the both termini. A 1.5 kb HindⅢ fragment of pCHU352 was cloned to Escherichia coli vector and it could be subcloned by exonucleaseⅢ deletion or restriction enzyme digestion. The sequences of this fragment were compared with the plasmid isolated from R. solani pathogenic strain CHU341.
URI: http://hdl.handle.net/11455/21654
Appears in Collections:生命科學系所

文件中的檔案:

取得全文請前往華藝線上圖書館



Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.