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Single-Cell Phenotypic Screening in Inverse Metabolic Engineering

  • A. E. VasdekisEmail author
  • G. Stephanopoulos
Chapter
  • 1.7k Downloads

Abstract

Contrary to classical approaches centering on debottlenecking flux-limiting steps in a metabolic pathway, inverse metabolic engineering (IME) aims at identifying and modulating all gene factors that contribute to an optimal phenotype. Within IME, mutant libraries are generated and screened in order to select mutants with the desired phenotype. The screening process is traditionally performed using microtiter well plates, a laborious and expensive process of limited throughput. Here, we review emerging screening methods that address these throughput and cost-effectiveness shortcomings, but also operate at the single-cell level. We discuss the importance of single-cell analyses in IME and detail two specific single-cell screening approaches: the first is fluorescence-activated cell sorting for phenotypic discrimination based on cytosolic or cell-membrane-bound products. The second is droplet microfluidics for screening of cells capable of overproducing secreted products or overconsuming substrates, properties that require confinement to isolate mutants with specific secretory phenotypes.

Keywords

Metabolic engineering Single-cell analysis Microfluidics Flow cytometry 

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Copyright information

© Springer International Publishing Switzerland 2016

Authors and Affiliations

  1. 1.Department of PhysicsUniversity of IdahoMoscowUSA
  2. 2.Department of Chemical EngineeringMassachusetts Institute of TechnologyCambridgeUSA

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