实验基础

This chapter outlines the basic requirements for finding and exploring sequences of interest in public databases, such as GenBank. As such, it is not aimed at experienced sequencers, for whom this will be “second nature,” but at the many clinical bacteriologists who rarely have need of DNA sequen ...

Comparative genomics is a science in its infancy. It has been driven by a huge increase in freely available genome-sequence data, and the development of computer techniques to allow whole-genome sequence analyses. Other approaches, which use hybridization as a method for comparing the gene ...

Many microbial genome sequences have been determined, and more new genome projects are ongoing. Shotgun sequencing of randomly cloned short pieces of genomic DNA can provide a simple way of determining whole genome sequences. This process requires sequencing of many fragments, compil ...

Living organisms are defined by the genes they possess. Control of expression of this gene set, both temporally and in response to the environment, determines whether an organism can survive changing conditions and can compete for the resources it needs to reproduce. Bacteria are no exceptio ...

The plethora of data now available from bacterial genome sequencing has opened a wealth of new research opportunities. Many of these have been reviewed in preceding chapters. Genomics alone, however, cannot capture a biological snapshot from an organism at a given point in time. The genome its ...

Microbiology has entered the postgenomic era and it is clear that bacterial typing should aim to be based on analysis of complete genomes. Although complete genome sequencing for epidemiological typing remains unrealistic for the present, microarrays that provide information on ge ...

Diagnostic medical bacteriology consists of two main components: identification and typing. Molecular biology has the potential to revolutionize the way in which diagnostic tests are delivered in order to optimize the care of infected patients, whether they are in hospital or in the com ...

Nucleic acid amplification technologies (NAATs) represent powerful tools in clinical microbiology, particularly in areas where traditional culture-based methods alone prove insufficient. A notable advantage is in reducing the time from taking samples to reporting result ...

This chapter describes two key strategies for the discovery of new antibacterial agents and illustrates the critical role played by genomics in each. The first approach is genomic target-based screening. Comparative genomics and bioinformatics are used to identify novel, selective ...

Genome sequencing, the determination of the complete complement of DNA in an organism, is revolutionizing all aspects of the biological sciences. Genome sequences make available for scientific scrutiny the complete genetic capacity of an organism. With respect to microbes, this mea ...

Microbial pathogens possess a repertoire of virulence determinants that make unique contributions to bacterial fitness during infection. In this chapter, we focus on the recent progress and adaptations of signature-tagged mutagenesis (STM) by PCR instead of hybridization. This ...

Recent advances in gene-amplification technology and molecular phylogenetics have provided the means of detecting and classifying bacteria directly from their natural habitats without the need for culture. These techniques have revolutionized environmental microbio ...

Isolates of bacterial species that are indistinguishable in genotype are assigned as a clone, with the implication that they are descended from the same recent ancestor. Clones are difficult to define with precision since bacteria are not truly asexual, and recombinational replaceme ...

Bacterial taxonomy comprises systematics (theory of classification), nomenclature (formal process of naming), and identification. There are two basic approaches to classification. Similarities may be derived between microorganisms by numerical taxonomic methods bas ...

Methods for isolation of small plasmids (usually cloning vehicles) from genetically characterized strains of Enterobacteriaceae (Escherichia coli and Salmonella) are well established (1–3). This chapter seeks to complement them by describing reliable basic methods for det ...

This chapter describes a simple method of making DNA probes with a nonisotopic label. The development of this type of new technology has, for example, created novel ways of detecting and identifying pathogenic microorganisms, which has led to a re-evaluation of methodological approaches ...

Antibiotic resistance is not an absolute property. Even within a single species, strains can exhibit different degrees of antibiotic resistance. When a bacterium is tested for resistance, it should be compared, quantitatively with a standard. This standard may be the resistance of a wild- ...

Plasmids are extra-chromosomal molecules of deoxyribonucleic acid (DNA) capable of autonomous replication. Such molecules have been identified in many bacterial genera and usually exist as covalently closed circular (CCC) molecules. Plasmids range in size from less than one meg ...

Traditional methods for the identification of some bacterial species can be time consuming and often necessitate the isolation of pure cultures before further characterization may be undertaken. Advances in molecular biology have allowed the identification of bacterial spec ...

Polymerase chain reaction (PCR) can be used to identify microorganisms in at least two basic ways. The first method depends on a knowledge of DNA sequence unique to the organism under study and provides a specific means of identifying that organism. This method is discussed elsewhere in this volu ...