TraDIS as a tool for accelerated lab-based evolution
Lead Research Organisation:
University of Birmingham
Department Name: Sch of Biosciences
Abstract
The use of long term laboratory experiments coupled with whole genome sequencing is currently proving a valuable tool for understanding the relationship between genotype and phenotype. Previous studies in our lab have shown that in replicated long term evolution at low pH, using E. coli as a model organism, mutations arise repeatedly in the same key regulator genes, and we have shown using strain construction and competition experiments that these are responsible for contributing the most to fitness of the evolved strains. Often, though not always, we were able to show that these were loss of function mutations. This project will study evolution of stress resistance under laboratory conditions in more detail by investigating further the contributions of individual genes to the fitness of the evolved strains and determining the mechanisms by which they act. One major issue with long term evolution is the length of time taken to conduct the experiment. We will therefore look at the potential of using high throughput methods for speeding up the generation and analysis of data from lab-based evolution experiments. If this approach is successful we will look at the roles of different parameters to see how robust the results are to parameter variation including the pH, the specific acid used, the media used, the strain genotype, the dilution rate, etc. This will enable us to link our data to more realistic "real world" scenarios and will be useful for informing future studies using lab-based evolution, for example for more efficient strain engineering or for studying emergence of resistance traits that might be important in organisms that cause food spoilage or infection.
Studentship Projects
Project Reference | Relationship | Related To | Start | End | Student Name |
---|---|---|---|---|---|
BB/M01116X/1 | 30/09/2015 | 31/03/2024 | |||
1790881 | Studentship | BB/M01116X/1 | 02/10/2016 | 25/01/2021 | Mathew Milner |
Description | Comparison of Essential genomes of Escherichia Coli using TraDIS |
Organisation | National University of Singapore |
Country | Singapore |
Sector | Academic/University |
PI Contribution | Construction of a high density UTI89 transposon library and subsequent analysis. Analysis of both MG1655 and ST131 transposon libraries. Comparison of all three transposon libraries for an essential genome comparison. Comparison of the essential genome of two separately created transposon libraries in UTI89 to ask the question is there lab specific variation between the construction of libraries |
Collaborator Contribution | National University of Singapore. Provision of UPEC strain UTI89 and creation and sequencing of one UTI89 Transposon library. University of Queensland. Discussion and assistance with the creation of scripts and analysis. |
Impact | N/A still undergoing analysis. |
Start Year | 2019 |
Description | Comparison of Essential genomes of Escherichia Coli using TraDIS |
Organisation | University of Birmingham |
Department | College of Life and Environmental Sciences |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | Construction of a high density UTI89 transposon library and subsequent analysis. Analysis of both MG1655 and ST131 transposon libraries. Comparison of all three transposon libraries for an essential genome comparison. Comparison of the essential genome of two separately created transposon libraries in UTI89 to ask the question is there lab specific variation between the construction of libraries |
Collaborator Contribution | National University of Singapore. Provision of UPEC strain UTI89 and creation and sequencing of one UTI89 Transposon library. University of Queensland. Discussion and assistance with the creation of scripts and analysis. |
Impact | N/A still undergoing analysis. |
Start Year | 2019 |
Description | Comparison of Essential genomes of Escherichia Coli using TraDIS |
Organisation | University of Queensland |
Department | Institute for Molecular Bioscience |
Country | Australia |
Sector | Academic/University |
PI Contribution | Construction of a high density UTI89 transposon library and subsequent analysis. Analysis of both MG1655 and ST131 transposon libraries. Comparison of all three transposon libraries for an essential genome comparison. Comparison of the essential genome of two separately created transposon libraries in UTI89 to ask the question is there lab specific variation between the construction of libraries |
Collaborator Contribution | National University of Singapore. Provision of UPEC strain UTI89 and creation and sequencing of one UTI89 Transposon library. University of Queensland. Discussion and assistance with the creation of scripts and analysis. |
Impact | N/A still undergoing analysis. |
Start Year | 2019 |
Description | Identification of genes required for, or involved in resistance to, a novel antimicrobial compound |
Organisation | University of Liverpool |
Country | United Kingdom |
Sector | Academic/University |
PI Contribution | Used methods arising from the BBSRC award on "Towards predictive biology: using stress responses in a bacterial pathogen to link molecular state to phenotype". Compound was provided by a company (Auspherix). We used traDIS methods with different concentrations of the compound to identify (a) genes which when mutated led to reduced growth at sub-mic concentrations and (b) genes which when mutated led to enhanced growth at sub-mic and greater than mic concentrations. Work was done by BBSRC-funded student whose project arose directly from the initial BBSRC award. |
Collaborator Contribution | Provided lab space, compound, and bioinformatics expertise. Research was funded by University of Liverpool spinout company. |
Impact | None to date but publication is expected as a specific finding has been made. Work is ongoing in developing and refining methods to link traDIS and whole genome sequence data from lab-evolved organisms with a view to enabling rapid prediction of key genes involved in novel resistance to antimicrobials. |
Start Year | 2018 |