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DNAase footprinting a simple method for the detection of protein-DNA binding specificity

机译:DNase足迹检测蛋白质-DNA结合特异性的简单方法

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A method for studying the sequence-specific binding of proteins to DNA is described. The technique is a simple conjoining of the Maxam-Gilbert DNA-sequencing method and the technique of DNAase-protected fragment isolation. Fragments of a 5′ end-labelled, double-stranded DNA segment, partially degraded by DNAase in the presence and absence of the binding protein, are visualized by eletrophoresis and autoradiography alongside the base-specific reaction products of the Maxam-Gilbert sequencing method. It is then possible to see the protective “footprint” of the binding protein on the DNA sequence. The binding of lac repressor to lac operator is visualized by “footprinting” as an example. Equilibrium estimates indicate that 10-fold sequence-specificity (differential binding constant) could be studied easily using this technique.
机译:描述了研究蛋白质序列特异性结合至DNA的方法。该技术是Maxam-Gilbert DNA测序方法的简单连合,以及DNA酶保护的片段分离技术。在存在和不存在结合蛋白的存在和不存在的情况下,通过DNA酶部分降解的5'末端标记的双链DNA段的片段,通过Eletrophoresis和Autoradopraphy与MaxAm-Gilbert测序方法的基本特异性反应产物进行了可视化。然后可以在DNA序列上看到结合蛋白的保护性“足迹”。 Lac压缩机与LAC操作者的结合以“足迹”为例而可视化。平衡估计表明可以使用该技术容易地研究10倍序列特异性(差动结合常数)。

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