The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study of Drosophila-colonizing Orbaceae.

Autor:	Elston KM; Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA., Phillips LE; Department of Integrative Biology, The University of Texas at Austin, Austin, TX, 78712, USA., Leonard SP; Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA.; Department of Integrative Biology, The University of Texas at Austin, Austin, TX, 78712, USA., Young E; Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA., Holley JC; Department of Integrative Biology, The University of Texas at Austin, Austin, TX, 78712, USA.; Freshman Research Initiative, The University of Texas at Austin, Austin, TX, 78712, USA., Ahsanullah T; Freshman Research Initiative, The University of Texas at Austin, Austin, TX, 78712, USA., McReynolds B; Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA., Moran NA; Department of Integrative Biology, The University of Texas at Austin, Austin, TX, 78712, USA., Barrick JE; Department of Molecular Biosciences, The University of Texas at Austin, Austin, TX, 78712, USA. jbarrick@cm.utexas.edu.
Jazyk:	angličtina
Zdroj:	ISME communications [ISME Commun] 2023 May 24; Vol. 3 (1), pp. 49. Date of Electronic Publication: 2023 May 24.
DOI:	10.1038/s43705-023-00255-3
Abstrakt:	Toolkits of plasmids and genetic parts streamline the process of assembling DNA constructs and engineering microbes. Many of these kits were designed with specific industrial or laboratory microbes in mind. For researchers interested in non-model microbial systems, it is often unclear which tools and techniques will function in newly isolated strains. To address this challenge, we designed the Pathfinder toolkit for quickly determining the compatibility of a bacterium with different plasmid components. Pathfinder plasmids combine three different broad-host-range origins of replication with multiple antibiotic resistance cassettes and reporters, so that sets of parts can be rapidly screened through multiplex conjugation. We first tested these plasmids in Escherichia coli, a strain of Sodalis praecaptivus that colonizes insects, and a Rosenbergiella isolate from leafhoppers. Then, we used the Pathfinder plasmids to engineer previously unstudied bacteria from the family Orbaceae that were isolated from several fly species. Engineered Orbaceae strains were able to colonize Drosophila melanogaster and could be visualized in fly guts. Orbaceae are common and abundant in the guts of wild-caught flies but have not been included in laboratory studies of how the Drosophila microbiome affects fly health. Thus, this work provides foundational genetic tools for studying microbial ecology and host-associated microbes, including bacteria that are a key constituent of the gut microbiome of a model insect species. (© 2023. The Author(s).)
Databáze:	MEDLINE
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