Showing papers by "Douglas B. Kell published in 2001"

A functional genomics strategy that uses metabolome data to reveal the phenotype of silent mutations

Abstract: A large proportion of the 6,000 genes present in the genome of Saccharomyces cerevisiae, and of those sequenced in other organisms, encode proteins of unknown function. Many of these genes are "silent," that is, they show no overt phenotype, in terms of growth rate or other fluxes, when they are deleted from the genome. We demonstrate how the intracellular concentrations of metabolites can reveal phenotypes for proteins active in metabolic regulation. Quantification of the change of several metabolite concentrations relative to the concentration change of one selected metabolite can reveal the site of action, in the metabolic network, of a silent gene. In the same way, comprehensive analyses of metabolite concentrations in mutants, providing "metabolic snapshots," can reveal functions when snapshots from strains deleted for unstudied genes are compared to those deleted for known genes. This approach to functional analysis, using comparative metabolomics, we call FANCY—an abbreviation for functional analysis by co-responses in yeast.

Discrimination of Aerobic Endospore-forming Bacteria via Electrospray-Ionization Mass Spectrometry of Whole Cell Suspensions

Abstract: Direct injection electrospray ionization mass spectrometry (ESI-MS) without prior analyte separation was investigated for the analysis of whole cell suspensions of bacteria. Thirty-six strains of aerobic endospore-forming bacteria, consisting of six Bacillus species and one Brevibacillus species, were studied. ESI was performed in the positive ion mode on the bacterial suspensions. Several peaks in the range of 250−1500 m/z were observed to contribute to variations in the spectral information among the species. Application of cluster analysis to the spectral data showed that this ESI-MS technique was capable of discriminating strains of the species B. subtilis. This investigation demonstrates the feasibility of measuring liquid samples with minimal sample preparation that can be useful for discrimination at the subspecies level. A change in the cone potential in the electrospray ion source was found to influence the spectral information of representative strains of all of the seven species tested. This ha...

Genomic computing. Explanatory analysis of plant expression profiling data using machine learning

Abstract: “Actually, the orgy of fact extraction in which everybody is currently engaged has, like most consumer economies, accumulated a vast debt. This is a debt of theory, and some of us are soon going to have an exciting time paying it back—with interest, I hope.” —Sydney Brenner, In Theory ,

Combining inductive logic programming, active learning and robotics to discover the function of genes

Abstract: The paper is addressed to AI workers with an interest in biomolecular genetics and also to biomolecular geneticists interested in what AI tools may do for them. The authors are engaged in a collaborative enterprise aimed at partially automating some aspects of scientific work. These aspects include the processes of forming hypotheses, devising trials to discriminate between these competing hypotheses, physically performing these trials and then using the results of these trials to converge upon an accurate hypothesis. As a potential component of the reasoning carried out by an "artificial scientist" this paper describes ASE-Progol, an Active Learning system which uses Inductive Logic Programming to construct hypothesised first-order theories and uses a CART-like algorithm to select trials for eliminating ILP derived hypotheses. In simulated yeast growth tests ASE-Progol was used to rediscover how genes participate in the aromatic amino acid pathway of Saccharomyces cerevisiae. The cost of the chemicals consumed in converging upon a hypothesis with an accuracy of around 88% was reduced by five orders of magnitude when trials were selected by ASE-Progol rather than being sampled at random. While the naive strategy of always choosing the cheapest trial from the set of candidate trials led to lower cumulative costs than ASE-Progol, both the naive strategy and the random strategy took significantly longer to converge upon a final hypothesis than ASE-Progol. For example to reach an accuracy of 80%, ASE-Progol required 4 days while random sampling required 6 days and the naive strategy required 10 days.

Chemometric criteria for the characterisation of Italian Protected Denomination of Origin (DOP) olive oils from their metabolic profiles

Abstract: This study was conducted and carried out as a consequence of the European directives n°2081/92 and n°2037/93 in which regulations for the protection of denomination of origin of food commodities were established. The research work was intended to investigate if monovarietal oils may be differentiated by their basic chemical composition. A positive outcome of this pilot study would hopefully permit extension to the characterisation of Italian DOP (Protected Denomination of Origin) olive oils. In the olive, long-chain alcohols, triterpenes and fatty acids, formed in distinct biosynthetic compartments, provide characteristic compositional data of an olive cultivar. The three classes of compounds were qualitatively and quantitatively determined by GC in six Italian olive cultivars, Coratina and Provenzale from Puglia, Frantoio and Moraiolo from Toscana, Bosana from Sardegna and Dritta from Abruzzo. Basic statistics and multivariate methods were first applied to the GC data of each class of compounds and then to the complete set of data with the aim of obtaining a clear discrimination of the cultivars. When Principal Components Analysis (PCA) was applied to the whole set of data consisting of alcohols, triterpenes and acids, the PCA revealed that the compounds (variables) that gave the better class distinction were: cycloartenol for Coratina, acids C20:0, C17:0, C18:0 for Dritta, citrostadienol for Frantoio and β-sitosterol for Moraiolo. Bosana and Provenzale correlated with erythrodiol and uvaol. A correct assignment of each oil sample to its monovarietal group was obtained.

MEG (Model Extender for Gepasi): a program for the modelling of complex, heterogeneous, cellular systems.

Abstract: Mendes, P., Kell, D. B. (2001). MEG (Model Extender for Gepasi): a program for the modelling of complex, heterogeneous, cellular systems. Bioinformatics, 17, (3), 288-290

Characterization of an autostimulatory substance produced by Escherichia coli

Abstract: The recovery of dilute populations of stationary phase cells of Escherichia coli was studied using an automatic growth analyser. The addition of 30% supernatant from 2-d-old stationary phase cells of the organism reproducibly shortened the apparent lag times by 22–57·5%, depending on the age of the inoculum. True lag times, as determined by colony counts, of stationary phase cells were reduced by supernatant addition by 41–62%. The growth-stimulating substance was characterized and partly purified from supernatants: the active material was shown to be dialysable, heat-stable, acid- and alkali-stable and protease-resistant. Extraction with ethyl acetate or ion-exchange resins was not successful, but the active material could be quantitatively extracted with ethanol after saturation with salt. It is concluded that the active substance is a small, non-proteinaceous, non-ionic organic molecule. Separation of extracts by HPLC indicated that the stimulatory substance is weakly hydrophobic and has retention times similar to those of uracil. So far, however, the exact chemical identity of the active substance has not been elucidated.

Developing a logical model of yeast metabolism

Abstract: With the completion of the sequencing of genomes of increasing numbers of organisms, the focus of biology is moving to determining the role of these genes (functional genomics). To this end it is useful to view the cell as a biochemical machine: it consumes simple molecules to manufacture more complex ones by chaining together biochemical reactions into long sequences referred to as em metabolic pathways. Such metabolic pathways are not linear but often interesect to form complex networks. Genes play a fundamental role in these networks by providing the information to synthesise the enzymes that catalyse biochemical reactions. Although developing a complete model of metabolism is of fundamental importance to biology and medicine, the size and complexity of the network has proven beyond the capacity of human reasoning. This paper presents the first results of the Robot Scientist research programme that aims to automatically discover the function of genes in the metabolism of the yeast em Saccharomyces cerevisiae. Results include: (1) the first logical model of metabolism;(2) a method to predict phenotype by deductive inference; and (3) a method to infer reactions and gene function by aductive inference. We describe the em in vivo experimental set-up which will allow these em in silico predictions to be automatically tested by a laboratory robot.

Histometrics: improvement of the dynamic range of fluorescently stained proteins resolved in electrophoretic gels using hyperspectral imaging.

Abstract: Most image-based analyses, using absorbance or fluorescence of the spatial distribution of identifiable structures in complex biological systems, use only a very small number of dimensions of possible spectral data for the generation and interpretation of the image. We here extend the concepts of hyperspectral imaging, being developed in remote sensing, into analytical biotechnology. The massive volume of information contained in hyperspectral spectroscopic images requires multivariate analysis in order to extract the chemical and spatial information contained within the data. We here describe the use of multivariate statistical methods to map and quantify common protein staining fluorophores (SYPRO Red, Orange and Tangerine) in electrophoretic gels. Specifically, we find (a) that the 'background' underpinning limits of detection is due more to proteins that have not migrated properly than to impurities or to ineffective destaining, (b) the detailed mechanisms of staining of SYPRO red and orange are apparently not identical, and in particular (c) that these methods can provide two orders of magnitude improvement in the detection limit per pixel, to levels well below the limit observable optically.

UNIT 11.3 Estimation of Microbial Viability Using Flow Cytometry

Abstract: For microorganisms in particular, viability is a term that is difficult to define and a state consequently difficult to measure. The traditional (and gold-standard) usage equates viability and culturability (i.e., the ability to multiply), but the process of determining culturability is often too slow. Flow cytometry provides the opportunity to make rapid and quantitative measurements of dye uptake in large numbers of cells, and we can therefore exploit the flow cytometric approach to evaluate so-called viability stains and to develop protocols for more routine assessments of microbial viability. This unit is primarily commentary, but several basic protocols have been included to ensure that users have a firm basis for attempting these reasonably difficult assays on traditional flow cytometer instruments. What is clear is that each assay must be carefully validated with the particular microorganism of interest before being applied in any research, clinical, or service form.

Spectral analysis via supervised genetic search with application-specific mutations

Abstract: We present a method in which a genetic algorithm is used to optimise an expression in order to provide a supervised method for interpretation of the infrared analytical spectra of complex biological samples. The aim is to produce a model that can predict the value of a measurand of interest, such as the concentration of a particular chemical constituent, from a complex infrared spectrum of biological material. The method we describe is in some ways analogous to genetic programming but it more readily allows the output expression to be constrained in complexity and permits its general form to be specified by the user, thereby enhancing its explanatory ability. The quasi-continuous properties of optical spectra are exploited by mutations that explore spectral regions adjacent to selected variables, and provide adaptive averaging of spectral regions so as to provide selective optimisation of the tradeoff between spectral resolution and signal-to-noise ratio.

Electrode with protective coating

Abstract: A metallic electrode (10) having at least one surface thereof provided with a protective coating (12), which protective coating (12) is substantially electrically transparent and comprises at least one copolymer comprising at least a first polymer moiety attaching at least a second polymer moiety to the surface, the at least second polymer moiety extending from the surface so as to inhibit contamination of the surface by at least one foreign substance. The electrode is particularly suitable for use in apparatus for analysing and/or monitoring biological material using non-linear dielectric spectroscopic techniques.

Apparatus and a method for solving problems

Abstract: A processor arrangement (2, 6, 11) is operable to provide an initial population of individuals each comprising a plurality of structural elements each comprising a plurality of functional components defining operations to be performed on at least one input to the functional component and then to: (i) activate each individual to cause the operations defined by the functional components of that individual to be performed to produce a result; (ii) determine from each result a fitness of the corresponding individual for providing a mechanism or program for providing a solution to the problem; (iii) produce a new generation of individuals by causing an evolutionary process to occur involving at least one structural element and at least one functional component; and then to repeat (i) to (iii) until either an individual of the current generation represents a suitable way, structure, mechanism or program for providing a solution to the problem or a predetermined number of generations have been evolved.