Chromatic Bacteria v.2-A Himar1 Transposon-Based Delivery Vector to Extend the Host Range of a Toolbox to Fluorescently Tag Bacteria

Mitja N P Remus-Emsermann,Christian Stocks,Rudolf O Schlechter

doi:10.3390/bacteria1010006

Mitja N P Remus-Emsermann, Christian Stocks + Show 1 more

Open Access

https://doi.org/10.3390/bacteria1010006

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Abstract

A recent publication described the construction and utility of a comprehensive “Chromatic Bacteria” toolbox containing a set of genetic tools that allows for fluorescently tagging a variety of Proteobacteria. In an effort to expand the range of bacteria taggable with the Chromatic Bacteria toolbox, a series of Himar1 transposon vectors was constructed to mediate insertion of fluorescent protein and antibiotic resistant genes. The Himar1 transposon was chosen as it is known to function in a wide range of bacterial species. To test the suitability of the new Himar1 Chromatic Bacteria plasmid derivatives, conjugations were attempted on recently isolated non-model organisms. Although we were unsuccessful in delivering the plasmids into Gram-positive bacterial isolates, we successfully modified previously recalcitrant isolates to the first set of the Chromatic Bacteria toolbox, such as Sphingomonas sp. Leaf357 and Acidovorax sp. Leaf84. This manuscript reports on the currently available plasmids and transposition success in different bacteria.

Highlights

We described the construction of a comprehensive “Chromatic Bacteria”
In this study we describe the construction and utility of Himar1-based suicide vectors that allow the fluorescent tagging of bacteria
We constructed a total of 23 plasmid vectors carrying different combinations of fluorescent proteins ranging from cyan to near-infrared fluorescence and antibiotic resistances including chloramphenicol, chloramphenicol and gentamicin; chloramphenicol and kanamycin; and chloramphenicol, kanamycin, and erythromycin (Figure 1, Table 1)

Summary

Introduction

Toolbox containing a set of plasmids, denoted the pMRE series, to fluorescently tag a wide range of bacterial isolates [1]. These plasmid tools were well received by the community and, at the time of writing this article, have been ordered more than 400 times from the nonprofit plasmid distribution platform addgene.org in the last 3 years alone It is possible to study them at the micrometer resolution. This is in stark contrast with the meta-omic research which has driven microbiology and microbial ecology during the last decade. Fluorescent tagging allows the study of bacteria at single cell resolution, which gives insight into biofilm formation, bacteria–bacteria and bacteria–host interactions [1]

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