Enhanced yeast one-hybrid assays for high-throughput gene-centered regulatory network mapping

UMMS Affiliation

Program in Gene Function and Expression; Program in Molecular Medicine; Program in Systems Biology; Department of Biochemistry and Molecular Pharmacology

Publication Date


Document Type



Two-Hybrid System Techniques; High-Throughput Screening Assays; Gene Regulatory Networks; Protein Interaction Mapping; Transcription Factors


Genetics and Genomics


A major challenge in systems biology is to understand the gene regulatory networks that drive development, physiology and pathology. Interactions between transcription factors and regulatory genomic regions provide the first level of gene control. Gateway-compatible yeast one-hybrid (Y1H) assays present a convenient method to identify and characterize the repertoire of transcription factors that can bind a DNA sequence of interest. To delineate genome-scale regulatory networks, however, large sets of DNA fragments need to be processed at high throughput and high coverage. Here we present enhanced Y1H (eY1H) assays that use a robotic mating platform with a set of improved Y1H reagents and automated readout quantification. We demonstrate that eY1H assays provide excellent coverage and identify interacting transcription factors for multiple DNA fragments in a short time. eY1H assays will be an important tool for mapping gene regulatory networks in Caenorhabditis elegans and other model organisms as well as in humans.

DOI of Published Version



Nat Methods. 2011 Oct 30;8(12):1059-64. doi: 10.1038/nmeth.1748. Link to article on publisher's site

Journal/Book/Conference Title

Nature methods

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Link to Article in PubMed

PubMed ID