Matrix and Steiner-triple-system smart pooling assays for high-performance transcription regulatory network mapping.
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.
Vermeirssen
Vanessa
V
Program in Gene Function and Expression and Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
Deplancke
Bart
B
Barrasa
M Inmaculada
MI
Reece-Hoyes
John S
JS
Arda
H Efsun
HE
Grove
Christian A
CA
Martinez
Natalia J
NJ
Sequerra
Reynaldo
R
Doucette-Stamm
Lynn
L
Brent
Michael R
MR
Walhout
Albertha J M
AJ
D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques