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題 名 | 具有轉錄活性之魚腥藻CH1蛋白粗抽萃取液=The Crude Extract from Anabaena sp. CH1 has the Transcriptional Activity |
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作 者 | 柴御清; 陳伯中; 陳良築; | 書刊名 | 中州學報 |
卷 期 | 18 2003.12[民92.12] |
頁 次 | 頁253-264 |
分類號 | 379.2 |
關鍵詞 | 魚腥藻; 生體外轉錄; RNA聚合脢; Anabaena; In vitro transcription; RNA polymerase; |
語 文 | 中文(Chinese) |
中文摘要 | 本研究利用polyethleneimine及ammonium sulfate等蛋白純化步廓,自臺灣水稻田中分離到的魚腥藻CH1品系中,大量製備其有轉錄活性的蛋白粗抽萃取液。在進行生體外轉錄反應分析之後,發現此魚腥藻蛋白蛋可以辨認菠菜葉綠體rbcL(ribulose-l.5-bisphosphate carboxylase/oxygenase large subunit)基因起動子,也可有效辨認大腸桿菌的終止子,但與葉綠體RNA聚合脢不同的是,魚腥藻RNA聚合脢萃取液對環狀及線狀的DNA模板均有轉錄的能力,而且轉錄的活性會受rifampicin抑制。 進一步利用聚合脢連鎖反應,篩選出魚腥藻CH1品系之rbcLS基因及其起動子,並將CH1品系 rbcLS基因起動子,殖入具有終止子之載體中,做為DNA模板,配合CH1蛋白萃取液進行生體外轉錄反應時,可轉錄出約780nt大小的專一性RNA產物;而菠菜葉綠體RNA聚合脢萃取液,則無法辨認此基因起動子。 |
英文摘要 | An active RNA polymerase crude extract from the Taiwan isolated cyanobacterium Anabaena sp. CH1 could be purified with polyethyeneimine and ammonium sulfate precipitation. This RNA polymerase extract could recognize the promoter of spinach chloroplast rbcL gene and the rbcLS gene from the Anabena strain CH1 and the terminator from the E. coli amino acid attenuator. In vitro transcription analysis indicated that the CH1 RNA polymerase crude extract could transcribe both supercoiled and linearized DNA template but the spinach chloroplast RNA polymerase crude extract could not transcribe linearized DNA template and they are sensitive to the antibiotic, rifampicin. The promoter region of rbcLS gene that isolated by polymerase chain reaction from CH1 chromosome and construct to the expression vector with the terminator from the E. coli amino acid attenuator for assay the crude extract transcriptional activity. While the CH1 RNA polymerase extract could recognize both the promoters of CH1 rbcLS gene and spinach chloroplast rbcL gene. the spinach chloroplast RNA polymerase could only recogize the spinach promoter but not the CH1 promoter. |
本系統中英文摘要資訊取自各篇刊載內容。