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Lookup NU author(s): Valeria Verrone,
Professor Anil Wipat
This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).
© 2021 by the authors. Licensee MDPI, Basel, Switzerland.Bacillus subtilis conjugative plasmid pLS20 uses a quorum-sensing mechanism to control expression levels of its conjugation genes, involving the repressor RcopLS20, the anti-repressor Rap-pLS20, and the signaling peptide Phr*pLS20. In previous studies, artificial overexpression of rappLS20 in the donor cells was shown to enhance conjugation efficiency. However, we found that the overex-pression of rappLS20 led to various phenotypic traits, including cell aggregation and death, which might have affected the correct determination of the conjugation efficiency when determined by colony formation assay. In the current study, conjugation efficiencies were determined under different conditions using a two-color fluorescence-activated flow cytometry method and measuring a single-round of pLS20-mediated transfer of a mobilizable plasmid. Under standard conditions, the conjugation efficiency obtained by fluorescence-activated flow cytometry was 23-fold higher than that obtained by colony formation. Furthermore, the efficiency difference increased to 45-fold when rappLS20 was overexpressed.
Author(s): Mori K, Verrone V, Amatsu R, Fukui K, Meijer WJJ, Ishikawa S, Wipat A, Yoshida K-I
Publication type: Article
Publication status: Published
Print publication date: 01/09/2021
Online publication date: 10/09/2021
Acceptance date: 08/09/2021
Date deposited: 22/09/2021
ISSN (electronic): 2076-2607
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