Current DTProg Projects

2017/2018


Student Christopher McCusker
Project Title Bio-production of natural herbicides in support of sustainable agriculture
Supervisor Katalin Kovacs, Elaine O'Reilly, Alex Conradie
School Life Sciences (34%), Chemistry (33%), Engineering (33%)

Project Description

SBRC Nottingham is one of three UK centres created by the BBSRC/EPSRC in 2014 and has received £14.3M in funding for a 5 year period. The proposed project will contribute towards generating new intellectual understanding and knowledge relating to metabolic engineering, enzyme engineering, computation biology systems and synthetic biology and its applications in sustainable production of natural herbicides from waste gasses using gas fermenting microbes. 

With a growing world population, maximising crop yield and minimising weed growth becomes ever more critical. Consequently, farmers turn to herbicides to boost crop yields. Given increasing weed resistance to current commercial herbicides, there is an urgent need for diversification of herbicide types, towards more environmentally friendly herbicides using more sustainable production processes. 
Chemosynthesis of a broader range of herbicides is possible, but relies on petrochemical feedstocks and may not be techno-economically feasible. A more innovative solution is a biosynthetic route through microbial fermentation using low-cost, abundant 1-carbon (C1) gases as the feedstock. In aid of sustainable agriculture as a UN development goal, the project will optimise a biochemical pathway through enzyme engineering, computation biology, systems and synthetic biology approaches. 

This 4-year PhD project is part of a University-funded Doctoral Training Programme in Synthetic Biology and associated with Nottingham’s new BBSRC/EPSRC Synthetic Biology Research Centre. This project aims to produce natural herbicides in a C1 metabolizing host in aid of sustainable agriculture as a UN development goal. A retrosynthesis algorithm will be utilised to generate a portfolio of novel, synthetic biochemical pathways from central metabolites to the target compounds. Students will also benefit from a diverse range of training opportunities, including specialist workshops, lectures and seminars, as well as participation in Nottingham’s yearly BBSRC DTP Spring School event. This project will be supervised by Dr Katalin Kovacs, Dr Elaine O’Reilly and Prof Alex Conradie. 

 
Student Claudio Tomi Andrino
Project Title Monitoring bacterial metabolism using global 13C-based metabolic flux analysis combined with LC-MS-based absolute quantification.
Supervisor Dong-Hyun Kim, Dave Barrett, John King, Klaus Winzer
School Pharmacy (50%), Mathematics (40%), Life Sciences (10%)

Project Description

The University of Nottingham seeks an enthusiastic PhD candidate to work on an exciting multi-disciplinary project that brings together the techniques of synthetic biology, analytical science and systems biology. 

This 4-year PhD project is part of a University-funded Doctoral Training Programme in Synthetic Biology and associated with Nottingham’s new BBSRC/EPSRC Synthetic Biology Research Centre. Students will benefit from a training in a range of widely applicable techniques, including through specialist workshops, lectures and seminars, as well as participation in Nottingham’s yearly BBSRC DTP Spring School event. This studentship will be based in the School of Pharmacy with collaborative links to the School of Life Sciences and the School of Mathematical Sciences. 

The project will focus on understanding bacterial cell metabolism under specific growth conditions and predicting metabolic phenotypes after genetic and/or environmental perturbations. Experimental work will involve the development and application of metabolic flux analysis based on LC-MS based metabolomics data. Quantitative models will built based on generated data in order to monitor the metabolism of a variety of bacterial species. The overall aim is to integrate metabolomics data into the systematic development of metabolic engineering strategies.

 
Student Francois Seys
Project Title Multiplex Genome Editing in Acetogens using CRISPR-based systems
Supervisor Nigel Minton, Ed Bolt
School Life Sciences (100%)

Project Description

CRISPR technology has become the mainstay of genome editing in all living organisms and is now synonymous with Synthetic Biology. Thanks to our international collaboration (BBSRC China partnership award) with the Key Laboratory of Synthetic Biology in Shanghai, we have made significant progress in the deployment of CRISRP/Cas9 for gene knock-in within our principle SBRC anaerobic chassis, Clostridium autoethanogenum [1, 2, 3]. It has revolutionised the speed with which we can make single mutants.  

The aim of the studentship is to maintain our lead in acetogen research by building on the developments we have made with CRISPR technology to explore the possibility of the simultaneous creation of multiple mutants in our anaerobic C1 chassis.  This will be achieved by both the derivation/ optimisation of CRISPRi approaches, while at the same time both exploring the possibility of: (i) introducing Non-Homologous End Joining (NHEJ) capability into acetogens, and; (ii) the use of Target-AID technology.

The output will be the development of systems with potentially wide application across the SBRC, while at the same time maintaining our lead in acetogen C1 technologies.

 
Student Amaury Montarnal
Project Title Engineering Escherichia coli for using carbon monoxide as the sole carbon and energy source.
Supervisor Philippe Soucaille, Naglis Malys, Minyeong Yoo
School Life Sciences (100%)

Project Description

SBRC Nottingham is one of three UK centres created by the BBSRC/EPSRC in 2014 and has received £14.3M in funding for a 5-year period. The proposed project will contribute towards generating new intellectual understanding and knowledge relating to metabolic engineering, enzyme engineering, computation biology, systems and synthetic biology and its applications in developing a metabolically engineered E. coli strain growing on carbon monoxide as the sole carbon and energy source. 

E. coli is industrially used for the production of bulk chemicals like amino acids (lysine, Threonene, tryptophane…) and diols (1, 3 popanediol, 1, 4 butanediol, 1, 2 propanediol…). However, due to the feedstocks cost, the production of a lot of bulk chemicals is currently not economical. 

Developing microorganism that could use cheap carbon source like carbon monoxide would largely expand the range of bulk chemicals that could be economically produced by E. coli. The project will aim to create a synthetic pathway for the use of carbon monoxide by E. coli as the sole carbon and energy source through computation biology, enzyme engineering, systems and synthetic biology approaches. 

This 4-year PhD project is part of a University-funded Doctoral Training Programme in Synthetic Biology and associated with Nottingham’s new BBSRC/EPSRC Synthetic Biology Research Centre. Students will also benefit from a diverse range of training opportunities, including specialist workshops, lectures and seminars, as well as participation in Nottingham’s yearly BBSRC DTP Spring School event.

 
Student Rudolf Hendriks
Project Title Synthetic Calvin Cycle for efficient CO2 fixation
Supervisor Philippe Soucaille, Katalin Kovacs, Minyeong Yoo
School Life Sciences (100%)

Project Description

SBRC Nottingham is one of three UK centres created by the BBSRC/EPSRC in 2014 and has received £14.3M in funding for a 5-year period. The proposed project will contribute towards generating new intellectual understanding and knowledge relating to metabolic engineering, enzyme engineering, computation biology, systems and synthetic biology and its applications in developing more energetically efficient CO2 fixation pathways in gas fermenting microbes and plants. 

Carbon fixation is the process by which CO2 is incorporated into organic compounds. In plants and many C1 fixing microorganisms, CO2 fixation is occurring through the Calvin–Benson cycle. This cycle which is the main pathway of CO2 fixation on earth is known to be energetically inefficient but so far nothing has been done to improve its energetic efficiency. 

Improving the energetic efficiency of the Calvin-Benson cycle would have a huge impact on the cost of production of bulk chemicals using C1 fixing microorganisms but would also improve growth and biomass yield of plants in the fields. The project will aim to create a synthetic Calvin-Benson cycle through computation biology, enzyme engineering, systems and synthetic biology approaches. 

This 4-year PhD project is part of a University-funded Doctoral Training Programme in Synthetic Biology and associated with Nottingham’s new BBSRC/EPSRC Synthetic Biology Research Centre. Students will also benefit from a diverse range of training opportunities, including specialist workshops, lectures and seminars, as well as participation in Nottingham’s yearly BBSRC DTP Spring School event.

 
Student Margaux Poulalier Delavelle
Project Title TBC
Supervisor  Nigel Minton
School  Life Sciences (100%)

Project Description

TBC

 

 

 

2016/2017

 
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Project Title  
Supervisor  
School  

Project Description

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2016/2017

Student

Project Title

Supervisor

School

Parisa Hamzavinejad Moghaddam

Sustainable Biocatalysis: Directed Evolution of Decarboxylase Enzyme

Neil Thomas,

Elaine O‘Reilly

Chemistry

Project description:

Synthetic Biology & Industrial Biotechnology offer a means of producing key organic molecules in a cleaner, greener and more sustainable way. In this research project, decarboxylase enzymes will be subjected to directed evolution (protein engineering/site-directed mutagenesis) in order to introduce new substrate and reaction specificity. The evolved enzymes will then be used either on their own, or as the initial catalyst in an artificial enzyme cascade to generate key high value intermediates for the pharmaceutical, chemical and food sectors. The effect of using a variety of non-aqueous reaction conditions on the evolved enzymes will also be examined. The PhD is one of a number funded by the Centre for Synthetic Biology at the University of Nottingham and you will join a cohort of students whose focus is on engineering proteins and metabolic pathways and will gain additional advanced training in synthetic biology. The work will be conducted in the Centre for Biomolecular Sciences which has state of the art facilities for protein manipulation and characterisation including X-ray crystallography, high field NMR, ITC, DSC, SPR and the School of Chemistry.

 

Student

Project Title

Supervisor

School

Joshua Luke Gascoyne 

Combinatorial engineering for production of short chain diols using synthetic metabolic pathways

Naglis Malys

Stephan Heeb

Life Sciences

Project description:

The PhD project will be aimed at engineering Cuprividus necator to produce diols such as 1,4-butanediol and 1,3-propanediol from carbohydrate sources and then ultimately using carbon dioxide and hydrogen as sole carbon and energy source. One of the project objectives will be to design and implement synthetic metabolic pathways for the production of these diols. The project will aim to develop and apply combinatorial transcriptional engineering methodologies for synthetic metabolic pathway optimisation. The genetically encoded metabolic switch that enables dynamic regulation of pathway expression by adapting metabolic activity to the changing environment will be sought. Bacterial microcompartments, sort of organelles composed of a protein shell with several functionally interlinked enzymes compartmentalised on the inside, will be another alternative that will provide solutions for diol production improvement through the increase in local metabolite flux and reduced intermediate toxicity to the cell. Development and optimisation of the gas fermentation process for bioproduction will play an important part of the project.

 

Student

Project Title

Supervisor

School

Nathan Dixon

Life Cycle Assessment for Responsible Research and Innovation in Synthetic Biology

John McKechnie

Engineering

Project description:

This project will integrate Life Cycle Assessment (LCA) and RRI methods to better understand the potential resource, environmental, and social implications of synthetic biology deployment. The integrated method will identify “hot spots” of high potential impact – e.g., feedstock requirements and procurement impacts, production impacts, product markets and social benefits and risks - and thereby inform ongoing technical research to reduce uneven or unintended consequences. The potential exists for social science to add value to LCA in a number of key ways, including accounting for social benefits and risks, incorporating diverse stakeholder perspectives and understanding ethical and value-driven drivers. The LCA-RRI framework will be applied to suitable case studies of relevance to the energy, chemicals, and/or pharmaceuticals sector.

 

Student

Project Title

Supervisor

School

Alessandro Orlandini    

Efficient conversion of a- into b-alanine via engineered phenylalanine/tyrosine mutase

Francesca Paradisi

Klaus Winzer

Chemistry

Life Sciences

Project description:

In the assembly of biosynthetic pathways for the production of 3-hydroxypropionic acid (3-HPO) and its derivatives, b-alanine is an ideal intermediate as it can be easily deaminated to produce 3-oxopropionic acid and, following further reduction, it will yield the 3-HPO.

However, in micro-organisms, the biosynthesis of b-alanine takes place via degradation pathways rather than by direct conversion of a-alanine, in fact a specific aminomutase capable of this biotransformation does not exist.

In this proposal we aim at targeting MIO-dependent aminomutases and evolve them through rational design as well as directed evolution to accept alanine as substrate. The generated mutants will then be incorporated into artificial pathways for the synthesis of 3-HPO. MIO-dependent aminomutases identified to date are only active on aromatic substrates (phenylalanine, PAM, and tyrosine, TAM) and no example of aliphatic MIO-dependent enzyme has been reported. This poses a very interesting challenge as evolution of this class of enzyme would definitely be more appealing due to their higher stability.

 

Student

Project Title

Supervisor

School

Joseph Ingram                                 

Tunable zinc responsive bacterial promoters for controlled gene expression

John Hobman

Dov Stekel

Phil Hill

Biosciences

Project description:

Zinc is an essential metal, required in ~30% of bacterial proteins, but is toxic at higher intracellular concentrations. Bacteria such as E. coli have evolved sophisticated zinc import and export systems controlled by transcription factors that repress the expression of genes encoding importer proteins (regulator Zur) or activate expression of zinc efflux (regulator ZntR). These regulators and the promoters they control represent a good example of fine tuning of cellular response to external zinc concentrations (1) and different Zur and ZntR regulated promoters have different affinities and transcription levels. We wish to study the levels of expression from engineered Zur and ZntR regulated promoters in response to zinc, so that a suite of promoters can be used to finely control gene expression in response to zinc levels in growth media. These promoters will be used to control gene expression in engineered bacteria using cheap zinc inducers and zinc chelators.

 

Student

Project Title

Supervisor

School

Mechelle Bennett          

Cellular-Redox Driven Fabrication of Biomimetic Polymers

Frankie Rawson

Phil Hill

Pharmacy

Biosciences

Project description:

The aim of this PhD will be to develop a new synthetic biology approach for the synthesis of biomimetic polymers, by genetic engineering of bacteria with iron reducing proteins to facilitate redox driven cell surface synthesis of polymers (FIG 1). The potential applications and therefore impact are broad in nature ranging from bacterial instructed routes in development of glycopolymer based drugs for treatment of disease, through to novel routes to manufacture biomimetic polymers to be used in tissue engineering and biodegradable plastics. Another interesting application is the ability to facilitate increases capture of cellular electricity in microbial fuel cells via directed growth of cell surface conducting polymers reducing our reliance on fossil fuels and petrochemicals.

 

Student

Project Title

Supervisor

School

Sean Craig                                          

Towards photosynthetic hydrogen production: regulation of electron supply to the cyanobacterial hydrogenase

Sam Bryan

Nigel Minton

Life Sciences

Project description:

Hydrogen has real potential as a clean renewable fuel, producing water on combustion. Solar powered bio-hydrogen has several advantages; it is relatively harmless to the organisms producing it and is easily separated from the growth media. Photoautotrophic microbes like cyanobacteria can utilize cheap and plentiful sources of carbon and electrons for growth and hydrogen production making them self-sustaining production vehicles. However, diverting a high proportion of reducing power to hydrogen production poses significant challenges, exacerbated considerably by uncertainties in how the hydrogenase interacts with the electron transport chain. We have shown that the hydrogenase has two distinct physiologically-controlled localization mechanisms that partition it within the thylakoids, either dispersed uniformly through the thylakoids or aggregated into discrete puncta (Burroughts et al., 2014). Crucially, electron supply and hydrogen production depend on physiologically controlled localization. Determination of the molecular basis for control of hydrogenase location could thus pave the way to engineering improved cyanobacterial cells for solar-powered bio-hydrogen production and will form the basis of this project. Specifically the project will aim to determine the signal transduction pathway that controls Hox localisation in Synechocystis sp. PCC6803 and determine the influence of Hox localisation on electron transport pathways and hydrogen production. The student will utilise a wide range of techniques including knockouts, pulldowns and flash photolysis. There will also be the opportunity to evolve the enzyme to improve productivity through error prone PCR. The project will involve multidisciplinary collaborations across Kiel University (Dr Jens Appel), Warwick University (Prof Nigel Burroughs) and UCL (Prof Paul Dalby).

 

 

Student

Project Title

Supervisor

School

Christian Gude                                 

Biosynthesis of value added chemicals by Cupriavidus necator H16

Kati Kovacs

Alex Conradie

Nigel Minton

Life Sciences

Project description:

Cupriavidus necator H16 is a Gram-negative, facultatively chemolithoautotrophic organism able to grow with organic substrates or CO2 / H2 under aerobic conditions. Its ability to grow on CO2 as sole carbon source makes it an ideal chassis organism for the sustainable production of platform chemicals from waste gases.

The aim of this project is to metabolically engineer Cupriavidus necator H16 for the sustainable production of value added chemicals from CO2 / H2. In particular, this project will evaluate producing chemical building blocks that are within three enzymatic steps from central metabolism, such building blocks having application as polymers or in sustainable agriculture.

 

2015/2016

Student

Project Title

Supervisor

School

Stelios Grigoriou

 

Flow Biocatalysis: a synthetically constructed bacterial bio-factory for the efficient synthesis of industrially important chiral amines

Dr Elaine O’Reilly

Dr Phil Hill

Prof John King

 

Chemistry

Project description:

The project aims to create a rationally designed, multicellular biocatalytic structure, exemplified using a chiral amine biofactory. The multi-bacterial structure will mimic flow chemistry processes, and be capable of supplying the enzyme substrates, producing the biocatalyst and trapping the high-value product. This self-sufficient bio-factory will enable cost-effective and sustainable routes to important amine motifs, will represent an attractive approach for the industrial synthesis of chiral amines and provide proof of concept for the creation of other multicellular biofactories.

Student

Project Title

Supervisor

School

Nathan Jones

 

DNA Replication of a Minimally-Sized Genome

Dr Thorsten Allers

Dr Jamie Twycross

SoLS

Project description:

The PhD project will use a synthetic biology approach to dissect the process of DNA replication in vivo and in vitro. DNA replication is the most fundamental task that proliferating cells must carry out, and begins at specific locations on the chromosome called origins. Archaea are the third domain of life, alongside eukaryotes and bacteria. Archaea are microbes renowned for living in extreme conditions such as acid pools and salt lakes. Haloferax volcanii comes from the Dead Sea and grows in saturated salt solutions. The enzymes that carry out DNA replication in archaea are strikingly similar to those used in eukaryotes. We have found that in the absence of origins, DNA replication depends instead on homologous recombination. The aim is to develop a simplified system for DNA replication based on homologous recombination. (1) To determine the minimum length for efficient replication by homologous recombination, we will construct large and small DNA molecules by genome engineering; (2) an inducible promoter will be used to regulate the expression of recombination genes, thereby facilitating the alternative mode of DNA replication; (3) a computational model will be developed to predict the rate of DNA replication, based on the length of recombining molecules and the rate of homologous recombination.

Student

Project Title

Supervisor

School

Alexandra Schindl

 

 

 

Experimental and insilico characterisation of halophilic Alcohol Dehydrogenase 2 in ionic liquids

Dr Anna Croft

Dr Mischa Zelzer

Dr Thorsten Allers

 

Engineering

Project description:

 

This project investigates systems comprising of the halophilic protein alcohol dehydrogenase 2 (HvADH2) from H. volcanii and a broad range of ionic liquids (ILs) with diverse physicochemical properties. Addition of ILs offers intriguing prospects for stabilising globular proteins and their enzymatic function against deactivating reaction channels of folding intermediates and misfolds promoting irreversible protein aggregation. We examine the impact of highly heterogeneous molecular electronic structures of 13 ionic liquids on protein structure and bio-catalytic activity. Our pursuit is to trace back the role of specific functional groups and their influence on the microenvironment of HvADH2.

Student

Project Title

Supervisor

School

Joshua Petch

 

Use of synthetic biology to fish out bacteria from sequential metabolic reactions

Dr Guiseppe Mantovani

Prof Miguel Camara

Dr Stephan Heeb

Pharmacy

Project description:

Large-scale selective sequestration of specific bacterial species and/or strains from mixed populations is a long term goal in a number of academic and industrial sectors, spanning from biotechnology to healthcare. In recent years a range of technologies, spanning from microfluidics to hydrogels and lipid-silica structures have been developed to this aim, although they are generally limited to the confinement of small bacterial populations in defined locations. Within the context of synthetic biology, microbial engineering has been playing an increasingly important role in the biotechnological production of high-value chemicals, including pharmaceuticals.

This multidisciplinary project aims to develop a platform technology to selectively sequester specific microorganisms from complex mixtures once they are no longer needed for a given biosynthetic pathway, overcoming the problem of non-desirable reactions. This will involve (i) assembly and characterisation functional surfaces displaying appropriate ligands with properties which minimise unspecific bacterial binding, (ii) engineering of suitable bacterial strains with inducible displays of membrane receptors, and (iii) bacterial binding/sequestration from mixed populations to functional surfaces.

Student

Project Title

Supervisor

School

Mattias Basle

 

Towards artificial metabolic pathways: design of non-natural imine reductases

Dr Anca Pordea

Dr Bas Winkler

Engineering

Project description:

The substrate and reaction scope of naturally occurring enzymes limits the production of high value, low molecular weight organic compounds by microorganisms. The availability of intracellular catalysts that facilitate non-natural transformations will therefore have enormous potential for the design of novel “intracellular retrosynthetic” pathways. To date, however, there are only few examples of chemical catalysts able to catalyse transformations in living organisms. To address this problem, we will design novel, non-natural imine reductases based on alcohol dehydrogenase scaffolds and evaluate their potential as building blocks for artificial metabolic pathways.

Student

Project Title

Supervisor

School

Erik Hanko

 

 

Switchable orthogonal gene control systems for metabolic engineering

Dr Naglis Malys

Prof Nigel P Minton

Life Science

Project description:

The project aims to develop a platform for switchable gene expression control systems that will effectively allow either to fine tune gene expression in rationally designed metabolic pathways increasing efficiency of production in biotechnologically important microorganisms or to switch-off completely production of those enzymes, which are no longer required increasing metabolic flux towards desired chemical or other biotechnological product. The project will involve several multidisciplinary components: (i) engineering, for developing logic circuits and designing orthogonal logic gates; (ii) computational, for modelling and predicting behaviour of the system; (iii) molecular biology and genetic engineering, for engineering of gene expression mechanisms in metabolically versatile bacteria with high biotechnological potential such as Cupriavidus necator. The project will also train the PhD applicant in microbiology, analytical biochemistry and wide spectrum of synthetic biology approaches. 

 

 

2014/2015

  Student

Project Title

Supervisor

School

Ms Vanisha Patel

High-throughput engineering and semi-directed evolution of bioreactor platform microbes

Dr Stephan Heeb

Dr Jamie Twycross, Dr Klaus Winzer

Computational  Sciences

Project description:

This synthetic biology project seeks to develop a principled approach to engineering microbes with minimal genomes adapted towards specified metabolic objectives. We will develop and combine novel wet lab (Vaud) and computational (Patel) techniques to create a high-throughput, broad host range pipeline capable of editing genes from target bacterial species to be transformed into designed chassis with the desired properties and enhanced exploitation potentials. Due to its relevance to industrial biotechnology, to its relatively close relatedness to Pseudomonas species for which we have extensive tools and expertise accumulated and to the innovative and challenging aspect of engineering it at the genomic level, we have selected Cupriavidus necator as the initial target organism.

Student

Project Title

Supervisor

School

Mr Christopher Stead

Methane to liquid transportation fuels using methanotrophic organisms

Dr Ying Zhang

Prof Nigel Minton

SoLS

Project description:

Methane is responsible for 10% of the world’s greenhouse gas emissions (on a CO2 equivalent basis), in part because it’s global-warming potential is twenty times greater than that of CO2 over a 100-year period. Using Synthetic Biology (SynBio) approaches, the studentship will be expected to design, implement artificial pathways and biological parts towards improving the performance of the recognised methanotrophic organisms (aerobic bacteria that utilise CH4 as the sole source of carbon and energy) that could potentially form the basis of a commercial process for the production of chemicals and fuels, particularly liquid transportation fuels.

Student

Project Title

Supervisor

School

Mr Paul Henry

Spectroscopic Insight into Microbial Fermentation

Prof Pete Licence

Chemistry

Project description:

The principle aim of this PhD project will be the development of a suite of in-situ spectroscopic techniques that will enable the real time analysis and instrumentation of live fermentation cultures in laboratory and production scale gas fermenters.  We will probe bulk product concentration and develop novel techniques to allow the concentration of intra-cellular metabolites (both quantitative and qualitative) to be established while the fermenter is under growth conditions without the need for invasive and potentially problematic sample extraction.  The main analysis tools will include a comprehensive suite of vibrational spectroscopies (particularly infra-red (including NIR) and Raman techniques, both employing multiple detector fibre optic probes.  To complement in-situ spectroscopy we will also use in-situ chromatographic techniques to validate quantitative data.

Student

Project Title

Supervisor

School

Ms Sophie Vaud

High-throughput engineering and semi-directed evolution of bioreactor platform microbes

Dr Stephan Heeb,

Dr Jamie Twycross Dr Klaus Winzer

SoLS

Project description:

This synthetic biology project seeks to develop a principled approach to engineering microbes with minimal genomes adapted towards specified metabolic objectives. We will develop and combine novel wet lab (Vaud) and computational (Patel) techniques to create a high-throughput, broad host range pipeline capable of editing genes from target bacterial species to be transformed into designed chassis with the desired properties and enhanced exploitation potentials. Due to its relevance to industrial biotechnology, to its relatively close relatedness to Pseudomonas species for which we have extensive tools and expertise accumulated and to the innovative and challenging aspect of engineering it at the genomic level, we have selected Cupriavidus necator as the initial target organism.