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Multiplex Genome Editing by Natural Transformation (MuGENT) for Synthetic Biology in Vibrio natriegens
journal contribution
posted on 2017-06-01, 00:00 authored by Triana N. Dalia, Chelsea A. Hayes, Sergey Stolyar, Christopher J. Marx, James B. McKinlay, Ankur B. DaliaVibrio natriegens has recently emerged as an alternative to Escherichia coli for molecular biology and biotechnology, but low-efficiency genetic
tools hamper its development. Here, we uncover how to induce natural
competence in V. natriegens and describe methods
for multiplex genome editing by natural transformation (MuGENT). MuGENT
promotes integration of multiple genome edits at high-efficiency on
unprecedented time scales. Also, this method allows for generating
highly complex mutant populations, which can be exploited for metabolic
engineering efforts. As a proof-of-concept, we attempted to enhance
production of the value added chemical poly-β-hydroxybutyrate
(PHB) in V. natriegens by targeting the expression
of nine genes involved in PHB biosynthesis via MuGENT.
Within 1 week, we isolated edited strains that produced ∼100
times more PHB than the parent isolate and ∼3.3 times more
than a rationally designed strain. Thus, the methods described here
should extend the utility of this species for diverse academic and
industrial applications.