AI Article Synopsis
- The study focuses on creating efficient genetic modification strategies for researchers to better understand gene functions in various applications.
- The researchers developed a system that includes three-gene cassettes (RPLK and RPCC) featuring antibiotic resistance for selection, a marker like mCherry, and a counterselection gene, achieving high transformation efficiency with fewer false positives.
- They successfully demonstrated this system by creating unmarked luminescent strains and mutants in different species, including the deletion of up to six genetic loci in a single strain.
Article Abstract
The development of simple and highly efficient strategies for genetic modifications is essential for postgenetic studies aimed at characterizing gene functions for various applications. We sought to develop a reliable system for species that allows for both unmarked and accumulation of multiple genetic modifications in a single strain. In this work, we developed and validated three-gene cassettes named RPLK and RPCC, comprising of an antibiotic resistance marker for positive selection, the phenotypic selection marker or mCherry, and the counterselection gene These cassettes can be transformed with high efficiency across the genus while significantly reducing the number of false positives compared with similar approaches. We exemplified the versatility and application of these systems by obtaining unmarked luminescent strains (knock-in) or mutants (knock-out) in different pathogenic and commensal species across the genus in addition to the cumulative deletion of six loci in a single strain of
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| http://dx.doi.org/10.1139/cjm-2022-0024 | DOI Listing |
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