Determination of the promoter strength in the mixed transcription system: promoters of lactose, tryplophan and ribosomal protein L10 operons fromExcherichia Coli

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Abstract
In vitrotranscription was performed in a single reaction mixture, which contained three species of truncated E.coliDNA template, each carrying one specific promoter, lacP (UV5), trpP orrpljp, and the transcripts of distinct sizes were analyzed by electrophoresis on the same gel. Using this “mixed transcription” system, the order of the promoter strength, i.e., the capacity to form stable open complex, was determined in the single-round transcription under the standard conditions (50mMNaCl and 37°C) to be lacP > trpP > rplJp the latter two promoters being 30˜40 and 5˜10% the strength of lacP, respectively. After the multiple-round transcription, however, the level of trp transcription was the lowest due to low cyclic-reaction rate but became the highest when another trp fragment containing the natural terminator was used as the template. The order of the transcription level also varied depending on the ionic strength and the reaction temperature and, as a result, lacP was no more the strongest under high salt concentration and at high temperature.