A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions
A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions
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DOI：
10.1073/pnas.110149297
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发表时间：
2000-06-20
影响因子：
11.1
通讯作者：
Pabo, CO
中科院分区：
文献类型：
--
作者：
Joung, JK;Ramm, EI;Pabo, CO
我们开发了一种细菌“双杂交”系统，可以很容易地从大于10(8)的文库中进行选择。我们的细菌系统可用于研究蛋白质-DNA或蛋白质-蛋白质的相互作用，与现有的基于酵母的单杂交和双杂交方法相比，它提供了许多潜在的显著优势。我们通过选择锌指变体(从一个大型随机文库中)与所需的DNA靶点紧密结合并特定地结合来测试我们的系统。我们的方法允许在单个选择步骤中分离特定序列的锌指。因此，它应该比噬菌体展示策略更快，后者通常需要多次浓缩/扩增周期。考虑到我们的基于细菌的选择系统可以处理的大型文库规模，这种方法应该为识别和优化蛋白质-DNA和蛋白质-蛋白质相互作用提供一个强大的工具。
We have developed a bacterial "two-hybrid" system that readily allows selection from libraries larger than 10(8) in size. Our bacterial system may be used to study either protein-DNA or protein-protein interactions, and it offers a number of potentially significant advantages over existing yeast-based one-hybrid and two-hybrid methods. We tested our system by selecting zinc finger variants (from a large randomized library) that bind tightly and specifically to desired DNA target sites. Our method allows sequence-specific zinc fingers to be isolated in a single selection step. and thus it should be more rapid than phage display strategies that typically require multiple enrichment/amplification cycles. Given the large library sizes our bacterial-based selection system can handle, this method should provide a powerful tool for identifying and optimizing protein-DNA and protein-protein interactions.