RT Journal Article
SR Electronic
T1 Establishing a cell-free <em>Vibrio natriegens</em> expression system
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 331645
DO 10.1101/331645
A1 Wiegand, Daniel J.
A1 Lee, Henry H.
A1 Ostrov, Nili
A1 Church, George M.
YR 2018
UL http://biorxiv.org/content/early/2018/05/26/331645.abstract
AB The fast growing bacterium Vibrio natriegens is an emerging microbial host for biotechnology. Harnessing its productive cellular components may offer a compelling platform for rapid protein production and prototyping of metabolic pathways or genetic circuits. Here, we report the development of a V. natriegens cell-free expression system. We devised a simplified crude extract preparation protocol and achieved &gt;260µg/mL of super-folder GFP in a small-scale batch reaction after three hours. Culturing conditions, including growth media and cell density, significantly affect translation kinetics and extract protein yield. We observed maximal protein yield at incubation temperatures of 26°C or 30°C, and show improved yield by tuning ions crucial for ribosomal stability. This work establishes an initial V. natriegens cell-free expression system, enables probing of V. natriegens biology, and will serve as a platform to accelerate metabolic engineering and synthetic biology applications.