BioRep-HGT: Bio-reporting horizontal gene transfer. An experimental tool that eliminates selection-dependency
The aim of this project is to develop bio-reporters that will report horizontal transfer of plasmids regardless of the presence of genes that enable selection. Such novel bio-reporters will advance research on horizontal gene transfer and thus bacterial evolution immensely.
Currently there is simply no efficient way of isolating plasmids without selection. This project suggests a risky, yet promising way of developing a research tool that will circumvent the selection bias. The reporter strains could both be used to isolate single plasmids and thoroughly characterize them, but could also be used in a high through-put manner, as FACS can be used to sort out many reporter stains with different plasmids. The plasmid DNA of these cells can then be isolated and sequenced, generating a plasmid meta-genome specific to the community.
BioRep-HGT: Bio-reporting horizontal gene transfer. An experimental tool that eliminates selection-dependency
Bacteria are the most abundant and diverse group of organisms on earth. They have critical roles in earth’s matter and element cycling, in addition to the health of all of its inhabitants. Yet, fundamental questions about bacterial evolution are still unanswered. Horizontal gene transfer (HGT), the transfer of genes between cells where the recipient is not the progeny (daughter cell) of the donor, is fundamental to bacterial evolution. Conjugative plasmids are genetic elements that promote ongoing and rapid HGT among bacteria. They transfer horizontally by conjugation, where a copy of the plasmid is transferred to a recipient upon cell-cell contact. Despite extensive evidence supporting the role of plasmids as key facilitators of HGT, and thus bacterial evolution, the understanding of what genes are transferred horizontally by plasmids, how often, and, from a Darwinian point of view, why, is presently very limited. This is due to critical experimental limitations.
Currently, selection for specific phenotypes encoded by plasmids is needed in order to isolate and characterize them. Researchers, therefore, typically look for transfer of genes that enable efficient selection, because it is otherwise notoriously difficult to pinpoint the very few community members that are subject to HGT. It is e.g. known that antibiotic resistance genes (ARGs), metal resistance genes and metabolic functions can be found on plasmids, but mainly because these are phenotypes that enable us to efficiently select in the laboratory. However, such selection-based approaches obviously give a biased impression of what genes are subject to HGT by plasmids, and it is clear that the majority of genes carried by plasmids have not been identified and thus not studied.
Here, I propose that bio-reporters can be constructed that report when conjugation of plasmids to the reporter bacterium occurs, regardless of what genes they carry, thus eliminating the need for selection.
Major gains
Currently there is simply no efficient way of isolating plasmids without selection. This proposal suggests a risky, yet promising way of developing a research tool that will circumvent the selection bias. The reporter strains could both be used to isolate single plasmids and thoroughly characterize them, but could also be used in a high through-put manner, as FACS can be used to sort out many reporter stains with different plasmids. The plasmid DNA of these cells can then be isolated and sequenced, generating a plasmid meta-genome specific to the community. Another important gain is that frequencies of plasmid transfer in the communities could be assessed – currently such important information is not attainable.
My research focuses on how bacteria interact, the molecular biology behind interactions, and how these social interactions evolve. Basically, the molecular biology and evolution of conflict, manipulation, sharing, and dependencies among bacteria.
I am especially interested in the role horizontal gene transfer – gene-sharing between bacteria – has in bacterial social evolution. The critical increase of antibiotic resistance among infectious bacteria, for example, is a striking and scary outcome of horizontal gene transfer. Sharing of genes is typically facilitated by mobile genetic elements and I studying how such elements interact with the rest of the host genome, in addition to how such interactions effect the social behavior of the host bacterium.
In order to experimentally address the hypothesis driven research briefly described above, an important part of my work is to develop molecular tools and engineer bacteria. The experimental research conducted by my group utilizes genome engineering (e.g. by CRISPR/Cas), synthetic biology, and fluorescence reporter gene technologies, FACS, CLSM, high-throughput sequencing, transcriptomics, and classical microbiology methodologies. We also use in vivo animal and plant models that are made available by collaborators with proper facilities.
Section of Microbiology
WHERE NO IDEAS ARE TOO small
Funded by:
Villum Experiment Grant from The Villum Foundation
Project title: BioRep-HGT: Bio-reporting horizontal gene transfer. An experimental tool that eliminates selection-dependency
Project period: September 2019 - August 2022
Contact
Assistant professor
Jonas Stenløkke Madsen
Universitetparken 15
Building 1, 1st floor
DK-2100 Copenhagen
Email: JSMadsen@bio.ku.dk