Expanding the genetic toolbox for the obligate human pathogenStreptococcus pyogenes Article Swipe
Nina Lautenschläger
,
Katja Schmidt
,
Carolin J. Schiffer
,
Thomas F. Wulff
,
Karin Hahnke
,
Knut Finstermeier
,
Moïse Mansour
,
Alexander Elsholz
,
Emmanuelle Charpentier
·
YOU?
·
· 2024
· Open Access
·
· DOI: https://doi.org/10.1101/2024.03.04.582890
YOU?
·
· 2024
· Open Access
·
· DOI: https://doi.org/10.1101/2024.03.04.582890
Genetic tools form the basis for the study of molecular mechanisms. Despite many recent advances in the field of genetic engineering in bacteria, genetic toolsets remain scarce for non-model organisms, such as the obligatory human pathogen Streptococcus pyogenes . In this study, we set out to develop a comprehensive set of plasmids, promoters and reporters for S. pyogenes . We present an expansion to the current genetic toolbox that comprises new replicative and site-specific integrative plasmids. Moreover, we established a collection of constitutive promoters with a wide variety of strengths as well as a set of novel inducible regulatory elements, including a zinc-inducible promoter, an erythromycin-inducible riboswitch and an IPTG-inducible promoter that outperform previously described inducible systems in terms of tightness and inducibility. In addition, we demonstrated the applicability of two codon-optimized fluorescent proteins, mNeongreen and mKate2, as reporters in S. pyogenes . For this, we adapted a novel chemically defined medium called RPMI4Spy. This medium showed a highly reduced autofluorescence compared to other growth media and allowed efficient signal detection in plate reader assays and fluorescence microscopy. Finally, we developed a plasmid-based system for genome engineering in S. pyogenes featuring the counterselection marker pheS *, which improved the generation of scarless gene deletions. This new toolbox simplifies previously laborious genetic manipulation procedures and lays the foundation for new methodologies to study gene functions in S. pyogenes, leading to a better understanding of its virulence mechanisms and physiology.
Related Topics
Concepts
Obligate
Streptococcus pyogenes
Toolbox
Human pathogen
Pathogen
Biology
Microbiology
Genetics
Gene
Computer science
Bacteria
Ecology
Programming language
Staphylococcus aureus
Metadata
- Type
- preprint
- Language
- en
- Landing Page
- https://doi.org/10.1101/2024.03.04.582890
- https://www.biorxiv.org/content/biorxiv/early/2024/03/04/2024.03.04.582890.full.pdf
- OA Status
- green
- References
- 100
- Related Works
- 10
- OpenAlex ID
- https://openalex.org/W4392383317
All OpenAlex metadata
Raw OpenAlex JSON
- OpenAlex ID
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https://openalex.org/W4392383317Canonical identifier for this work in OpenAlex
- DOI
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https://doi.org/10.1101/2024.03.04.582890Digital Object Identifier
- Title
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Expanding the genetic toolbox for the obligate human pathogenStreptococcus pyogenesWork title
- Type
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preprintOpenAlex work type
- Language
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enPrimary language
- Publication year
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2024Year of publication
- Publication date
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2024-03-04Full publication date if available
- Authors
-
Nina Lautenschläger, Katja Schmidt, Carolin J. Schiffer, Thomas F. Wulff, Karin Hahnke, Knut Finstermeier, Moïse Mansour, Alexander Elsholz, Emmanuelle CharpentierList of authors in order
- Landing page
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https://doi.org/10.1101/2024.03.04.582890Publisher landing page
- PDF URL
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https://www.biorxiv.org/content/biorxiv/early/2024/03/04/2024.03.04.582890.full.pdfDirect link to full text PDF
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YesWhether a free full text is available
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greenOpen access status per OpenAlex
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https://www.biorxiv.org/content/biorxiv/early/2024/03/04/2024.03.04.582890.full.pdfDirect OA link when available
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Obligate, Streptococcus pyogenes, Toolbox, Human pathogen, Pathogen, Biology, Microbiology, Genetics, Gene, Computer science, Bacteria, Ecology, Programming language, Staphylococcus aureusTop concepts (fields/topics) attached by OpenAlex
- Cited by
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0Total citation count in OpenAlex
- References (count)
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100Number of works referenced by this work
- Related works (count)
-
10Other works algorithmically related by OpenAlex
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