REGULATION OF RIBOSOMAL-RNA PROMOTERS WITH A SYNTHETIC LAC OPERATOR
REGULATION OF RIBOSOMAL-RNA PROMOTERS WITH A SYNTHETIC LAC OPERATOR
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DOI：
10.1073/pnas.81.22.6929
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发表时间：
1984-01-01
期刊：
影响因子：
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通讯作者：
HOLY, A
中科院分区：
文献类型：
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作者：
BROSIUS, J;HOLY, A
在克隆的大肠杆菌rrnB rRNA启动子P2启动子的起始点附近，顺向和反向插入一段长达21个碱基对的DNA片段，该DNA片段中含有一个中心的Lac操纵子序列。在Lac IQ菌株中，RNA合成在两个方向上都被有效地抑制，并被异丙基-β-D-硫代半乳糖苷诱导。当P2上游的rrnB启动子P1和合成的lac操纵子也存在时，转录抑制是不完整的。转录水平是在体内测量的，间接通过蛋白质(氯霉素乙酰转移酶)的表达，或直接通过稳定的RNA(E.coli4.5S RNA)的表达来测量，在一种简单的方法中，包括对未标记的大肠杆菌总RNA的凝胶电泳法。RrnB启动子的构建可以产生高水平的蛋白质表达和稳定的RNA表达。
A synthetic 21-base-pair long DNA fragment containing the central lac operator sequence was inserted near the initiation point of the cloned Escherichia coli rrnB rRNA promotor P2 in the natural and reverse orientation. RNA synthesis is efficiently repressed in both orientations in lac Iq strains and is induced with isopropyl .beta.-D-thiogalactoside. When the rrnB promotor P1 is also present, upstream from P2 and the synthetic lac operator, repression of transcription is incomplete. The levels of transcription were measured in vivo, indirectly by the expression of a protein (chloramphenicol acetyltransferase), or directly by the expression of a stable RNA (E. coli 4.5S RNA) in a simple assay involving gel electrophoresis of unlabeled total RNA from E. coli. The rrnB promotor constructions can produce high levels of protein expression as well as high levels of expression of stable RNA.