Matrix and Steiner-triple-system smart pooling assays for high-performance transcription regulatory network mapping

Program in Gene Function and Expression and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
Nature Methods (Impact Factor: 25.95). 09/2007; 4(8):659-64. DOI: 10.1038/nmeth1063
Source: PubMed

ABSTRACT Yeast one-hybrid (Y1H) assays provide a gene-centered method for the identification of interactions between gene promoters and regulatory transcription factors (TFs). To date, Y1H assays have involved library screens that are relatively expensive and laborious. We present two Y1H strategies that allow immediate prey identification: matrix assays that use an array of 755 individual Caenorhabditis elegans TFs, and smart-pool assays that use TF multiplexing. Both strategies simplify the Y1H pipeline and reduce the cost of protein-DNA interaction identification. We used a Steiner triple system (STS) to create smart pools of 4-25 TFs. Notably, we uniplexed a small number of highly connected TFs to allow efficient assay deconvolution. Both strategies outperform library screens in terms of coverage, confidence and throughput. These versatile strategies can be adapted both to TFs in other systems and, likely, to other biomolecules and assays as well.

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Available from: Vanessa Vermeirssen, Nov 14, 2014
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    Genetic Epidemiology 12/2013; 37(8). DOI:10.1002/gepi.21769 · 2.95 Impact Factor
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    • "Mating experiments were performed as described previously (Walhout et al, 2000; Vermeirssen et al, 2007). Briefly, the two 384-well AD-TF plates were transformed into the Ya1867 strain (a kind gift from John Reece-Hoyes and Marian Walhout, University of Massachusetts Medical School, USA). "
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