An Accessible Continuous-Culture Turbidostat for Pooled Analysis of Complex Libraries

Author:

McGeachy Anna M.,Meacham Zuriah A.,Ingolia Nicholas T.ORCID

Abstract

ABSTRACTWe present an accessible, robust continuous-culture turbidostat system that enables biologists to generate and phenotypically analyze highly complex libraries in yeast and bacteria. Our system has many applications in genomics and systems biology; here, we demonstrate three of these uses. We first measure how the growth rate of budding yeast responds to limiting nitrogen at steady state and in a dynamically varying environment. We also demonstrate the direct selection of a diverse, genome-scale protein fusion library in liquid culture. Finally, we perform a comprehensive mutational analysis of the essential gene RPL28 in budding yeast, mapping sequence constraints on its wild-type function and delineating the binding site of the drug cycloheximide through resistance mutations. Our system can be constructed and operated with no specialized skills or equipment and applied to study genome-wide mutant pools and diverse libraries of sequence variants under well-defined growth conditions.

Publisher

Cold Spring Harbor Laboratory

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