Christopher Edward French

The University of Edinburgh | UoE · School of Biological Sciences

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly.Each Modular cloning toolkit uses different vectors that can be compatible or not with other systems. And the vectors in each toolkit have advantages and disadavantges based on what their intended use. JUMP vector...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly.Each Modular cloning toolkit uses different vectors that can be compatible or not with other systems. And the vectors in each toolkit have advantages and disadavantges based on what their intended use. JUMP vector...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly. Despite being reliable and automation-friendly, each standard uses a specific set of vectors, requiring researchers to generate new toolkits for novel hosts and cloning applications. JUMP vectors (1) combine the...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly.Each Modular cloning toolkit uses different vectors that can be compatible or not with other systems. And the vectors in each toolkit have advantages and disadavantges based on what their intended use. JUMP vector...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly. Despite being reliable and automation-friendly, each standard uses a specific set of vectors, requiring researchers to generate new toolkits for novel hosts and cloning applications. JUMP vectors (1) combine the...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly.Each Modular cloning toolkit uses different vectors that can be compatible or not with other systems. And the vectors in each toolkit have advantages and disadavantges based on what their intended use. JUMP vector...

Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly.Each Modular cloning toolkit uses different vectors that can be compatible or not with other systems. And the vectors in each toolkit have advantages and disadavantges based on what their intended use. JUMP vector...

About Modular Cloning and JUMP: Modular cloning standards based on Golden Gate DNA assembly allow construction of complex DNA constructs over several rounds of assembly. Despite being reliable and automation-friendly, each standard uses a specific set of vectors, requiring researchers to generate new toolkits for novel hosts and cloning application...

Creating DNA constructs is a basic and fundamental step in molecular and synthetic biology. While prices for gene synthesis are decreasing, it is still more economical in most cases to assemble constructs from a library of components (Parts). Many methods for DNA assembly are available, but most require either a fixed and inflexible format for the...

Modular cloning standards based on Golden Gate DNA assembly allow for construction of complex DNA constructs over several rounds of assembly. Despite being reliable and automation-friendly, each standard uses a specific set of vectors, requiring researchers to generate new tool kits for novel hosts and cloning applications. JUMP vectors (Valenzuela...

In vivo logic gates have proven difficult to combine into larger devices. Our cell-based logic system, ParAlleL, decomposes a large circuit into a collection of small subcircuits working in parallel, each subcircuit responding to a different combination of inputs. A final global output is then generated by a combination of the responses. Using ParA...

Cell-free systems (CFS) are an emerging technology where cell extracts are used instead of living genetically-modified cells creating a completely in vitro reaction, thus, not presenting a "living prospect" applicable to current legal regulations. Here we show a versatile cell-free system based on the master survivalist bacteria Cupriavidus metalli...

In vivo logic gates have proven difficult to combine into larger devices. Our cell-based logic system, BioLogic, decomposes a large circuit into a collection of small subcircuits working in parallel, each subcircuit responding to a different combination of inputs. A final global output is then generated by combination of the responses. Using BioLog...

Green macroalgae represent an attractive source of renewable carbon. Conversion of green algal biomass to useful products requires depolymerisation of the cell wall polysaccharide ulvan. The enzymatic depolymerisation of ulvan is not fully understood and only few enzymes involved in this process have been reported. In this study the first model for...

Bacillus subtilis is a valuable industrial production platform for proteins, a bacterial model for cellular differentiation and its endospores have been proposed as a vehicle for vaccine delivery. As such B. subtilis is a major synthetic biology chassis but, unlike Escherichia coli , lacks a standardized toolbox for genetic manipulation. EcoFlex is...

Joining DNA sequences to create linear and circular constructs is a basic requirement in molecular biology. Here we describe PaperClip, a recently developed method, which enables assembly of multiple DNA sequences in one reaction in a combinatorial manner. In contrast to other homology-based multi-part assembly methods currently available, PaperCli...

Estimation of bacterial growth by rapid traditional methods such as spectrophometric measurements at 600nm (OD600) is not applicable for cultures containing insoluble particles in the growth media. Colony counts are the only suitable alternative but these are laborious and not high-throughput. The data presented in this article is related to the re...

INTRODUCTION Although much effort has been devoted to studying yeast in the past few decades, our understanding of this model organism is still limited. Rapidly developing DNA synthesis techniques have made a “build-to-understand” approach feasible to reengineer on the genome scale. Here, we report on the completion of a 770-kilobase synthetic yeas...

Delivery of DNA to cells and its subsequent integration into the host genome is a fundamental task in molecular biology, biotechnology and gene therapy. Here we describe an IP-free one-step method that enables stable genome integration into either prokaryotic or eukaryotic cells. A synthetic mariner transposon is generated by flanking a DNA sequenc...

Monitoring bacteria growth is an important technique required for many applications such as testing bacteria against compounds (e.g. drugs), evaluating bacteria composition in the environment (e.g. sewage and wastewater or food suspensions) and testing engineered bacteria for various functions (e.g. cellulose degradation). Traditionally, rapid esti...

Significance and impact of the study: Biofuels have been shown to be the best sustainable and alternative source of fuel to replace fossil fuels. Of the different types of feedstocks used for producing biofuels, lignocellulosic biomass is the most abundant. Converting this biomass to useful products has met with little success. Different approache...

Effective degradation of cellulose requires multiple classes of enzyme working together. However, naturally occurring cellulases with multiple catalytic domains seem to be rather rare in known cellulose-degrading organisms. A fusion protein made from Cellulomonas fimi exo- and endo- glucanases, Cex and CenA which improves breakdown of cellulose is...

The mechanism of antibacterial action of silver nanoparticles (AgNp) was investigated by employing a combination of microbiology and geochemical approaches to contribute to the realistic assessment of nanotoxicity. Our studies showed that suspending AgNp in media with different levels of chloride relevant to environmental conditions produced low le...

This BioBrick Foundation Request for Comments (BBF RFC 104) describes a new approach to multiple part DNA assembly – BrickClip, which does not require use of any restriction enzymes, nor cloning of the parts to specific donor and acceptor vectors. BrickClip allows assembly of up to six parts from existing parts collections, including the Registry o...

Synthetic biology allows the generation of complex recombinant systems using libraries of modular components. Two major near-market applications are whole-cell biosensors and biocatalysts for conversion of lignocellulosic biomass to biofuels and chemical feedstocks. Whole cell biosensors consist of cells genetically modified so that binding of a sp...

Ethanol is an important biofuel. Heterologous expression of Zymomonas mobilis pyruvate decarboxylase (Pdc) and alcohol dehydrogenase (AdhB) increases ethanol production in Escherichia coli. A fusion of PDC and ADH was generated and expressed in E. coli. The fusion enzyme was demonstrated to possess both activities. AdhB activity was significantly l...

Assembly of DNA ‘parts’ to create larger constructs is an essential enabling technique for bioengineering and synthetic biology. Here we describe a simple method, PaperClip, which allows flexible assembly of multiple DNA parts from currently existing libraries cloned in any vector. No restriction enzymes, mutagenesis of internal restriction sites,...

This BioBricks Foundation Request for Comments (BBF RFC) describes a novel approach, GenBrick, for the rapid assembly of multiple BioBrick RFC10-compatible parts in a single reaction, with completely flexible part order, without recloning or reamplification.

As described elsewhere in this volume, it is widely accepted that cellulosic biomass represents an enormous reservoir of renewable reduced carbon, and is the only feasible source for even partial replacement of our current use of fossil fuels as a source of material for transport fuels and chemical feedstocks. For decades, microbiologists and engin...

The increasing production and use of engineered nanoparticles, coupled with their demonstrated toxicity to different organisms, demands the development of a systematic understanding of how nanoparticle toxicity depends on important environmental parameters as well as surface properties of both cells and nanomaterials. We demonstrate that production...

In bacteria, the production of exopolysaccharides-polysaccharides secreted by the cells into their growth medium-is integral to the formation of aggregates and biofilms. These exopolysaccharides often form part of a matrix that holds the cells together. Investigating the bacterium Sinorhizobium meliloti, we found that a mutant that overproduces the...

Arsenic is a serious problem in groundwater in Bangladesh, West Bengal, and other regions, with up to 100 million people worldwide estimated to be at risk. Long term consumption leads to arsenicosis, skin lesions and cancers. There is a clear need for cheap, simple, non-toxic field test kits for routine monitoring of arsenic levels in drinking wate...

Arsenic contaminated groundwater is estimated to affect over 100 million people worldwide, with Bangladesh and West Bengal being among the worst affected regions. A simple, cheap, accurate and disposable device is required for arsenic field testing. We have previously described a novel biosensor for arsenic in which the output is a change in pH, wh...

The use of permeable pavement systems with integrated geothermal heat pumps for the treatment and recycling of urban runoff is novel and timely. This study assesses the efficiency of the combined technology for controlled indoor and uncontrolled outdoor experimental rigs. Water quality parameters such as biochemical oxygen demand, nutrients, total...

This Request for Comments (RFC) modifies the assembly standard for biological parts proposed in BBF RFC 10 by removing the NotI restriction site from the BioBrick Prefix and Suffix.

We study phase separation in suspensions of two unrelated species of rod-like bacteria, Escherichia coli and Sinorhizobium meliloti, induced by the addition of two different anionic polyelectrolytes, sodium polystyrene sulfonate or succinoglycan, the former being synthetic and the latter of natural origin. Comparison with the known behaviour of syn...

Arsenic is a serious problem in groundwater in Bangladesh, West Bengal, and other regions, with up to 100 million people worldwide estimated to be at risk. Long term consumption leads to arsenicosis, skin lesions and cancers. There is a clear need for cheap, simple, non-toxic field test kits for routine monitoring of arsenic levels in drinking wate...

To move our economy onto a sustainable basis, it is essential that we find a replacement for fossil carbon as a source of liquid fuels and chemical industry feedstocks. Lignocellulosic biomass, available in enormous quantities, is the only feasible replacement. Many micro-organisms are capable of rapid and efficient degradation of biomass, employin...

This BioBricks Foundation Request for Comments (BBF RFC) provides in- formation about the preparation and submission of BBF RFCs to The Bio- Bricks Foundation (BBF).

Plants are increasingly being employed to clean up environmental pollutants such as heavy metals; however, a major limitation of phytoremediation is the inability of plants to mineralize most organic pollutants. A key component of organic pollutants is halogenated aliphatic compounds that include 1,2-dichloroethane (1,2-DCA). Although plants lack t...

Poster presentation.

Hemerythrins are oxygen-binding proteins found in the body fluids and tissues of certain invertebrates. Oxygen is bound at a nonheme iron centre consisting of two oxo-bridged iron atoms bound to a characteristic set of conserved histidine: aspartate and glutamate residues with the motifs H-HxxxE-HxxxH-HxxxxD. It has recently been demonstrated bioch...

We sought to develop a whole-cell biosensor for the detection of arsenic in drinking water, a major problem in Bangladesh and West Bengal. In contrast to previously described systems, our biosensor would give a pH change as output, allowing simple detection with a pH electrode or pH indicator solution. We designed and modelled a system based on the...

This chapter summarises recent developments in whole cell biosensors for estimating toxicity and for the detection of pollutants in wastewater treatment plants. Biosensors of this sort started with the bioluminescence decay toxicity tests and it was realised that bioluminescence is a uniquely suited reporter system. Developments in molecular biolog...

The manufacture and improper disposal of explosives has resulted in a significant amount of land requiring remediation. The compound 2,4,6-trinitrotoluene (TNT) is the most persistent and toxic of the explosive pollutants with current treatment methods being energy intensive and costly. Bacterial enzymes such as pentaerythritol tetranitrate reducta...

There is major international concern over the wide-scale contamination of soil and associated ground water by persistent explosives residues. 2,4,6-Trinitrotoluene (TNT) is one of the most recalcitrant and toxic of all the military explosives. The lack of affordable and effective cleanup technologies for explosives contamination requires the develo...

Widespread contaimination of the environment by explosives resulting from the manufacture, disposal and testing of munitions is becoming a matter of increasing concern. Most explosives are considered to be a major hazard to biological systems due to their toxic and mutagenic effects. Interest on the bioremediation of land contaminated with explosiv...

Widespread contamination of the environment by explosives resulting from the manufacture, disposal and testing of munitions is becoming a matter of increasing concern. Most explosives axe considered to he a major hazard to biological systems due to their toxic and mutagenic effects. Interest on the bioremediation of land contaminated with explosive...

The discovery of widespread explosives contamination of soil, sediment, and groundwater has led to increasing concern about the persistence of the toxic compounds in the environment. The selective pressure of environmental pollution is clearly developing microorganisms that might be harnessed for explosives removal by biotechnological processes. Th...

We have applied the soluble pyridine nucleotide transhydrogenase ofPseudomonas fluorescens to a cell-free system for the regeneration of the nicotinamide cofactors NAD and NADP in the biological production of the important semisynthetic opiate drug hydromorphone. The original recombinant whole-cell system suffered from cofactor depletion resulting...

There is major international concern over the wide-scale contamination of soil and associated ground water by persistent explosives residues. 2,4,6-Trinitrotoluene (TNT) is one of the most recalcitrant and toxic of all the military explosives. The lack of affordable and effective cleanup technologies for explosives contamination requires the develo...

The gene encoding the soluble pyridine nucleotide transhydrogenase (STH) of Azotobacter vinelandii was cloned and sequenced. This is the third sth gene identified and further defines a new subfamily within the flavoprotein disulfide oxidoreductases. The three STHs identified all lack one of the redox active cysteines that are characteristic for thi...

Morphine dehydrogenase (MDH) of Pseudomonas putida M10 catalyses the NADP(+)-dependent oxidation of morphine and codeine to morphinone and codeinone. This enzyme forms the basis of a sensitive detection and assay method for heroin metabolites and a biotransformation process for production of hydromorphone and hydrocodone. To improve these processes...

Plants offer many advantages over bacteria as agents for bioremediation; however, they typically lack the degradative capabilities of specially selected bacterial strains. Transgenic plants expressing microbial degradative enzymes could combine the advantages of both systems. To investigate this possibility in the context of bioremediation of explo...

The udhA gene of Escherichia coli was cloned and expressed in E. coli and found to encode an enzyme with soluble pyridine nucleotide transhydrogenase activity. The N-terminal end of the enzyme contains the fingerprint motif of a dinucleotide binding domain, not present in published E. coli genome sequences due to a sequencing error. E. coli is here...

One of the major environmental problems facing the military establishment is the considerable amount of land and water that is contaminated with explosives, particularly TNT (2,4,6-trinitrotoluene). Explosives are found in the environment as a result of waste water produced by manufacturing plants and from the disposal of off-specification material...

Enterobacter cloacae PB2 was originally isolated on the basis of its ability to utilize nitrate esters, such as pentaerythritol tetranitrate (PETN) and glycerol trinitrate, as the sole nitrogen source for growth. The enzyme responsible is an NADPH-dependent reductase designated PETN reductase. E. cloacae PB2 was found to be capable of slow aerobic...

Pentaerythritol tetranitrate (PETN) reductase of Enterobacter cloacae PB2, a flavoprotein involved in the biodegradation of the explosive PETN, ethylene glycol dinitrate (EGDN) and glycerol trinitrate (GTN), was purified from an overexpressing strain of E. coli and crystallized at 293 K using the sitting-drop vapour-diffusion method. Diffraction da...

Morphinone reductase from Pseudomonas putida M10, a flavoprotein involved in the degradation of morphine alkaloids, was purified from an overexpressing strain of Escherichia coli and crystallized using the hanging-drop vapour-diffusion method. Diffraction data were collected to 2.5 A. The I-centred orthorhombic cell has a monomer in the asymmetric...

The gene encoding the soluble pyridine nucleotide transhydrogenase (STH) of Pseudomonas fluorescens was cloned and expressed in Escherichia coli. STH is related to the flavoprotein disulfide oxidoreductases but lacks one of the conserved redox-active cysteine residues. The gene is highly similar to an E. coli gene of unknown function.

Pentaerythritol tetranitrate reductase, which reductively liberates nitrite from nitrate esters, is related to old yellow enzyme. Pentaerythritol tetranitrate reductase follows a ping-pong mechanism with competitive substrate inhibition by NADPH, is strongly inhibited by steroids, and is capable of reducing the unsaturated bond of 2-cyclohexen-1-on...

A mixed microbial culture capable of metabolizing the explosive pentaerythritol tetranitrate (PETN) was obtained from soil enrichments under aerobic and nitrogen-limiting conditions. A strain of Enterobacter cloacae, designated PB2, was isolated from this culture and was found to use PETN as a sole source of nitrogen for growth. Growth yields sugge...

Morphinone reductase, produced by Pseudomonas putida M10, catalyses the NADH-dependent saturation of the carbon-carbon double bond of morphinone and codeinone, and is believed to be involved in the metabolism of morphine and codeine. The structural gene encoding morphinone reductase, designated morB, was cloned from Ps. putida M10 genomic DNA by th...

Semisynthetic derivatives of morphine and related alkaloids are in widespread clinical use. Due to the complexity of these molecules, however, chemical transformations are difficult to achieve in high yields. We recently identified the powerful analgesic hydromorphone as an intermediate in the metabolism of morphine by Pseudomonas putida M10. Here...

Semisynthetic opiates provide some of the most potent analgesic compounds currently in clinical use, the majority of which are synthesized from the naturally occurring alkaloids morphine, codeine and thebaine. The use of recombinant DNA technology to engineer pathways for the biological synthesis of semisynthetic opiate drugs could offer significan...

The NADH-dependent morphinone reductase from Pseudomonas putida M10 catalyses the reduction of morphinone and codeinone to hydromorphone and hydrocodone respectively. Morphinone reductase was purified from crude cell extracts to apparent homogeneity in a single affinity-chromatography step using Mimetic Yellow 2. The purified enzyme was a dimeric f...