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Microbiology Society logo Microbiology

Volume 171, Issue 4

Deletion of in removes autofluorescence in the excitation-emission range of cyan fluorescent protein Open Access

Abstract

Autofluorescence poses an impediment to fluorescence microscopy of biological samples. In the Gram-positive, soil-dwelling bacteria of the genus , sources of autofluorescence have not been examined systematically to date. Here, we show that the model organism for the genus, , shows autofluorescence in two of the commonly used fluorescence channels for visualizing cyan and green/yellow fluorescent proteins. We identify the source of autofluorescence in the cyan fluorescence channel as redox cofactor factor 420 (F) and target its synthesis to remove it. By deleting the () gene, which is a key biosynthetic gene for the production of F, we were able to create an autofluorescence-free strain in the cyan range of fluorescence excitation-emission. We demonstrate the usefulness of this strain by imaging the mTurquoise-tagged polar growth-related protein DivIVA and the cell division-related protein FtsZ in the deletion background. Using live-cell imaging to follow the dynamics of DivIVA and FtsZ, we demonstrate an improved signal-to-noise ratio in the mutant strain. We show that this strain can be a suitable tool for visualizing the localization of proteins in spp. and can facilitate the utilization of multi-colour imaging and fluorescence resonance energy transfer-based imaging.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): Actinomycetota , autofluorescence , F420 and Streptomyces
Funding
This study was supported by the:
  • Carl Tryggers Stiftelse för Vetenskaplig Forskning (Award CTS 19: 102)
    • Principle Award Recipient: KlasFlärdh
  • Vetenskapsrådet (Award 2019-04643)
    • Principle Award Recipient: KlasFlärdh
  • Kungliga Fysiografiska Sällskapet i Lund (Award 42451-2021)
    • Principle Award Recipient: ParminderSingh Mavi
  • Sven och Lilly Lawskis Fond för Naturvetenskaplig Forskning
    • Principle Award Recipient: ParminderSingh Mavi
© 2025 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
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2025-04-15
2025-04-23
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/content/journal/micro/10.1099/mic.0.001552
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Deletion of fbiC in Streptomyces venezuelae removes autofluorescence in the excitation-emission range of cyan fluorescent protein
Microbiology 171, 001552 (2025); https://doi.org/10.1099/mic.0.001552
/content/journal/micro/10.1099/mic.0.001552
/content/journal/micro/10.1099/mic.0.001552
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