Potassium permanganate as an in situ probe for B-Z and Z-Z junctions.
Potassium permanganate as an in situ probe for B-Z and Z-Z junctions.
复制标题
高锰酸钾作为 B-Z 和 Z-Z 连接处的原位探针。
DOI：
10.1093/nar/19.24.6943
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发表时间：
1991
影响因子：
14.9
通讯作者：
Amirhaeri,S
中科院分区：
文献类型：
--
作者：
Jiang,H;Zacharias,W;Amirhaeri,S
可应用于活细胞的DNA结构探针的可用性对于分析体内采用的不寻常DNA结构的生物学功能是必不可少的。我们已经开发了一种化学探针检测和定量的左手Z-DNA结构的重组质粒在生长E。大肠杆菌高锰酸钾选择性地与细胞中超冷质粒的B-Z或Z-Z连接区反应。位于这些连接处的限制性内切酶识别位点在用KMnO 4修饰后不被相应的核酸内切酶切割。这种切割的抑制允许测定细胞内克隆插入物的B-和Z-形式的相对量。我们已经成功地应用这种方法来监测Z-DNA形成的程度在E。大肠杆菌的生长期和突变的拓扑异构酶或促旋酶活性的函数。原则上，该测定法可用于任何在结构改变内部或附近含有限制性识别位点的不寻常DNA结构。它可以作为一个有用的工具来分析在体内DNA结构和基因调控事件之间的相关性。
The availability of DNA structural probes that can be applied to living cells is essential for the analysis of biological functions of unusual DNA structures adoptedin vivo. We have developed a chemical probe assay to detect and quantitate left-handed Z-DNA structures in recombinant plasmids in growingE. colicells. Potassium permanganate selectively reacts with B-Z or Z-Z junction regions in supercolled plasmids harbored in the cells. Restriction enzyme recognition sites located at these junctions are not cleaved by the corresponding endonuclease after modification with KMnO4. This inhibition of cleavage allows the determination of the relative amounts of B- and Z-forms of the cloned inserts Inside the cell. We have successfully applied this method to monitor the extent of Z-DNA formation inE. colias a function of the growth phase and mutated topoisomerase or gyrase activities. The assay can in principle be used for any unusual DNA structure that contains a restriction recognition site inside or near the structural alteration. It can be a useful tool to analyzeIn vivocorrelations between DNA structure and gene regulatory events.