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. 2019 Jun 20;9:8915. doi: 10.1038/s41598-019-45429-z

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Demonstration of a circularization-dependent promoter activity. (a) Different insertions of an aphA-3 gene together with a promoterless cat gene close to LJ or RJ of ICEHptfs4, or with an additional T-HP1395 terminator (T) upstream of the cat gene, were generated in strain P12, resulting in the indicated arrangements after selection for kanamycin or erythromycin resistance (see Materials and Methods for details). Subsequently, kanamycin- or erythromycin-resistant transformants were plated on chloramphenicol-containing media. A (+) sign indicates growth of colonies on chloramphenicol plates, and a (−) sign indicates no growth. (b) Genomic DNA preparations of chloramphenicol-selected colonies or non-selected (Kan/Erm) bacteria of mutant (7), or of P12 wild-type, were analysed by PCR with primer pair WS362/WS363 (refer to Fig. 3 for primer binding sites).