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  • Title: Construction of lycopene-overproducing E. coli strains by combining systematic and combinatorial gene knockout targets.
    Author: Alper H, Miyaoku K, Stephanopoulos G.
    Journal: Nat Biotechnol; 2005 May; 23(5):612-6. PubMed ID: 15821729.
    Abstract:
    Identification of genes that affect the product accumulation phenotype of recombinant strains is an important problem in industrial strain construction and a central tenet of metabolic engineering. We have used systematic (model-based) and combinatorial (transposon-based) methods to identify gene knockout targets that increase lycopene biosynthesis in strains of Escherichia coli. We show that these two search strategies yield two distinct gene sets, which affect product synthesis either through an increase in precursor availability or through (largely unknown) kinetic or regulatory mechanisms, respectively. Exhaustive exploration of all possible combinations of the above gene sets yielded a unique set of 64 knockout strains spanning the metabolic landscape of systematic and combinatorial gene knockout targets. This included a global maximum strain exhibiting an 8.5-fold product increase over recombinant K12 wild type and a twofold increase over the engineered parental strain. These results were further validated in controlled culture conditions.
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