A liquid handling platform for standardised quantification of cell-free enzymatic activity encoded by antimicrobial resistance genes
File(s)
Accepted version
Author(s)
Bergum, Molly
Martin, Bethany
Sutton, Mark
Moore, Simon
Type
Journal Article
Abstract
Antimicrobial resistance (AMR) is a growing global threat to human health, and rapid methods for characterising emerging antimicrobial resistance genes (ARGs) are needed. Here, we develop a semi-automated workflow using cell-free gene expression (CFE) systems to measure the activity of two ARGs encoded on plasmid DNA that produce rifampicin-inactivating and gentamicin-inactivating enzymes. We validated the use of a small benchtop Myra liquid handling system compared to manual pipetting, with no statistical differences observed. After optimising the pre-incubation time of ARGs and dispensing protocol, expression of aac(3)-IIa increased the half-maximal inhibition concentration (IC50) of gentamicin by over 150-fold, while arr-3 increased the IC50 of rifampicin by approximately 20-fold compared to controls. This methodology for rapid, semi-automated ARG characterisation offers a strategy to combat AMR by assessing novel ARGs identified through genomic surveillance or profiling activity of new or derivative antibiotics.
Date Acceptance
2026-08-19
Citation
Synthetic Biology
ISSN
1939-7267
Publisher
Oxford University Press
Journal / Book Title
Synthetic Biology
Copyright Statement
Copyright This paper is embargoed until publication. Once published the Version of Record (VoR) will be available on immediate open access.
License URL
Publication Status
Accepted