Gong Yunchen, Desveaux Darrell, Guttman David S, Lewis Jennifer D
Department of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada, M5S 3B2.
Centre for the Analysis of Genome Evolution and Function, University of Toronto, Toronto, ON, Canada, M5S 3B2.
Methods Mol Biol. 2017;1613:1-20. doi: 10.1007/978-1-4939-7027-8_1.
Yeast two-hybrid screens are a powerful approach to identify protein-protein interactions; however, they are typically limited in the number of interactions identified, and lack quantitative values to ascribe confidence scores to the interactions that are obtained. We have developed a high-throughput, quantitative, yeast two-hybrid screening approach coupled with next-generation sequencing. This strategy allows the identification of interacting proteins that are preferentially associated with a bait of interest, and helps eliminate nonspecific interacting proteins. The method is high-throughput, allowing many more baits to be tested and many more candidate interacting proteins to be identified. Quantitative data allows the interactors to be ascribed confidence scores based on their enrichment with particular baits, and can identify both common and rare interacting proteins.
酵母双杂交筛选是鉴定蛋白质-蛋白质相互作用的一种强大方法；然而，它们通常在鉴定出的相互作用数量上受到限制，并且缺乏定量值来为所获得的相互作用赋予置信度分数。我们开发了一种高通量、定量的酵母双杂交筛选方法，并结合了下一代测序技术。这种策略能够鉴定出优先与感兴趣的诱饵相关联的相互作用蛋白，并有助于消除非特异性相互作用蛋白。该方法具有高通量，能够测试更多的诱饵，并鉴定出更多的候选相互作用蛋白。定量数据允许根据相互作用蛋白与特定诱饵的富集情况为其赋予置信度分数，并且能够识别常见和罕见的相互作用蛋白。
