Shifted Transversal Design smart-pooling for high coverage interactome mapping.
"Smart-pooling," in which test reagents are multiplexed in a highly redundant manner, is a promising strategy for achieving high efficiency, sensitivity, and specificity in systems-level projects. However, previous applications relied on low redundancy designs that do not leverage the full potential of smart-pooling, and more powerful theoretical constructions, such as the Shifted Transversal Design (STD), lack experimental validation. Here we evaluate STD smart-pooling in yeast two-hybrid (Y2H) interactome mapping. We employed two STD designs and two established methods to perform ORFeome-wide Y2H screens with 12 baits. We found that STD pooling achieves similar levels of sensitivity and specificity as one-on-one array-based Y2H, while the costs and workloads are divided by three. The screening-sequencing approach is the most cost- and labor-efficient, yet STD identifies about twofold more interactions. Screening-sequencing remains an appropriate method for quickly producing low-coverage interactomes, while STD pooling appears as the method of choice for obtaining maps with higher coverage.
Xin
Xiaofeng
X
Banting and Best Department of Medical Research and Department of Molecular Genetics, Terrence Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario M5S 3E1, Canada.
Rual
Jean-François
JF
Hirozane-Kishikawa
Tomoko
T
Hill
David E
DE
Vidal
Marc
M
Boone
Charles
C
Thierry-Mieg
Nicolas
N
D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics*; D020869:Gene Expression Profiling; D020411:Oligonucleotide Array Sequence Analysis; D016366:Open Reading Frames / Q000235:genetics; D025941:Protein Interaction Mapping; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*; D012984:Software; D020798:Two-Hybrid System Techniques