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Curr Protoc Plant Biol ; 4(1): e20086, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-30742367

RESUMEN

Gene-centered yeast one-hybrid (Y1H) screens using arrayed genome-wide transcription factor (TF) clone collections provide a simple and effective strategy to identify TF-promoter interactions using a DNA fragment as bait. In an effort to improve the assay we recently developed a Y1H system that uses a cell surface Gaussia luciferase reporter (gLUC59). Compared to other available methods, this luciferase-based strategy requires a shorter processing time, enhances the throughput and improves result analysis of gene-centered Y1H screens. Here, we described the procedure to perform high-throughput screens using this novel strategy, which involves a protocol for mating two haploid yeast strains carrying an arrayed TF clone collection and a promoter::gLUC59 reporter, respectively, and a protocol for analyzing gLUC59 activity in the resulting diploid cells. © 2019 by John Wiley & Sons, Inc.


Asunto(s)
Genes Reporteros , Ensayos Analíticos de Alto Rendimiento/métodos , Luciferasas/metabolismo , Saccharomyces cerevisiae/metabolismo , Técnicas del Sistema de Dos Híbridos , Diploidia , Biblioteca de Genes , Mediciones Luminiscentes , Saccharomyces cerevisiae/citología , Factores de Transcripción/metabolismo
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