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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75562| 作者: | 陳景康 |
| 出版年 : | 1986 |
| 學位: | 碩士 |
| 摘要: | 水稻白葉枯病病原菌之RNA聚合?在其溶解性噬菌體Xp10感染後，會遺失其σ次單位．為了探討σ次單位遺失之原因，乃純化出RNA聚合?核?(core enzyme)，並以核?為抗原製備抗血清．以免疫沈澱法分析Xp10感染前後粗酵素萃取液中RNA聚合?組成，發現σ次單位會因Xp10的感染而降低對核?之吸附能力． 於純化過程中發現一個分子量為29,000的蛋白質ε，會與全?(holozyme)一起被純化出來．DEAE sephadex A 25管柱層析可將ε與全?分離．將ε與RNA聚合?重組後，發現可增進其轉錄活性，並可促進全?對Xp10 DNA的吸附能力．ε亦可於粗酵素萃取液中，以抗核?之抗血清沈澱出來．根據以上結果，認定ε為水稻白葉枯病病原菌RNA聚合?之次單位． 於實驗過程中亦發現核酸限制?XorII活性伴隨RNA聚合?出現．兩者間有無相關性，仍須進一步的研究． The σ subunit of Xanthomonas campestris pv. oryzae RNA polymerase was loss during bacteriophage Xp10 infection. To study the mechanism of σ loss, RNA polymerase from uninfected cells was purified to homogeneity by DNase treatment, ultracentrifugation, ammonium sulfate fractionation, Heparin sepharose 4B, Bio Rex 70, Bio gel A 1.5 m and DEAE sephadex A 25 chromatography. Both holozyme and core enzyme from RNA polymerase could be obtained by these procedures. Core enzyme was used to prepare antiserum for further study. Immunoprecipitations of anti-core antiserum vs. infected and uninfected crude extracts were performed and compared by SDS-PAGE followed by silver stain. The amount of σ subunit coprecipitated with core enzyme of infected crude extract was far less than that of uninfected crude extract. Such result suggested that σ subunit exhibited less binding ability to core enzyme after Xp10 infection. A 29,000 daltons protein named ε was found to copurify with holozyme. It also coprecipitated with anti-core antiserum. ε separated from holozyme through DEAE sephadex A 25 chromatography. After reconstitution with holozyme in vitro, the increase of transcription activity on Xp10 DNA and poly d(A-T) was observed. ε alond did not bind to Xp10 DNA, however, after binding with holozyme, the increase of binding efficiency to Xp10 DNA was detected. It is concluded that ε is a subunit of X. campestris pv. oryzae RNA polymerase. A restriction enzyme activity similar to XorII was detected. The role of thus enzyme activity in RNA polymerase is still unknown, it needs further investigation. |
| URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75562 |
| 全文授權: | 未授權 |
| 顯示於系所單位： | 植物科學研究所 |
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