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Single-step genome engineering in the bee gut symbiont Snodgrassella alvi

posted on 2024-06-11, 07:22 authored by The University of Texas at Austin
Honey bees are economically relevant pollinators experiencing population declines due to a number of threats. As in humans, the health of bees is influenced by their microbiome. The bacterium Snodgrassella alvi is a key member of the bee gut microbiome and has a role in excluding pathogens. Despite this importance, there are not currently any easy-to-use methods for modifying the S. alvi chromosome to study its genetics. To solve this problem, we developed a one-step procedure that uses electroporation and homologous recombination, which we term SnODIFY (Snodgrassella-specific One-step gene Deletion or Insertion to alter FunctionalitY). We used SnODIFY to create seven single-gene knockout mutants and recovered mutants for all constructs tested. Nearly all transformants had the designed genome modifications, indicating that SnODIFY is highly accurate. Mutant phenotypes were validated through knockout of Type 4 pilus genes, which led to reduced biofilm formation. We also demonstrate inserting heterologous sequences into the genome by integrating fluorescent protein-coding genes. Finally, we confirmed that genome modification is dependent on S. alvis endogenous RecA protein. Because it does not require expressing exogenous recombination machinery, SnODIFY is a straightforward, accurate, and lightweight method for genome editing in S. alvi. This workflow can be used to study the functions of S. alvi genes and to engineer this symbiont for applications, such as protecting honey bee health.


U.S. Department of Agriculture, 2023-67012-39356

National Science Foundation, IOS-2103208

National Institutes of Health, R35GM131738

U.S. Army Research Office, W911NF-20-1-0195


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National Center for Biotechnology Information

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  • Non-geospatial

ISO Topic Category

  • biota

National Agricultural Library Thesaurus terms

sequence analysis

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  • No

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  • Public

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It is recommended to cite the accession numbers that are assigned to data submissions, e.g. the GenBank, WGS or SRA accession numbers. If individual BioProjects need to be referenced, state that "The data have been deposited with links to BioProject accession number PRJNA1017617 in the NCBI BioProject database ("

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