实验基础

Site-specific recombinases such as the Saccharomyces cerevisiae Flp and the P1 phage Cre proteins have been increasingly used for the construction of unmarked deletions in bacteria. Both systems consist of an antibiotic resistance gene flanked by recognition sites in direct orient ...

The ability of bacteria to survive in a variety of different niches is due, in part, to their ability to respond and adapt to the environment. Extracellular signals are recognized by bacilli, and their responses are generally conducted at the transcript level. RNA polymerases recognize speci ...

A wide variety of inducible expression systems have been designed for Gram-negative bacteria, but adapting these systems to phylogenetically distinct species, such as mycobacteria, has proved notoriously difficult. Mycobacteria belong to a class of high G+C Gram-positive bacte ...

Elucidating the function of mycobacterial proteins, determining their contribution to virulence, and developing new vaccine candidates has been facilitated by systems permitting the heterologous expression of genes in mycobacteria. Mycobacterium bovis bacille Calme ...

The construction of allelic exchange mutants is one of the major strategies to decipher the function of a defined gene. In this chapter, protocols are described to perform allelic exchange in the mycobacterial species Mycobacterium smegmatis and Mycobacterium tuberculosis using th ...

The importance of plasmids for molecular research cannot be underestimated. These double-stranded DNA units that replicate independently of the chromosomal DNA are as valuable to bacterial geneticists as a carpenter’s hammer. Fortunately, today the mycobacterial research co ...

With the increased need for novel antimicrobials to improve the existing treatment for tuberculosis, to combat multidrug-resistant tuberculosis, and to address the presence of latent bacilli in a large population throughout the world, which can reactivate and cause active disease, ...

An agar dilution method for measuring minimum inhibitory concentrations (MICs) of Mycobacterium tuberculosis, based on the method of proportion, is described. Mycobacterium strains are grown on Middlebrook 7H10 (or 7H11) agar medium with twofold serially diluted drug concentr ...

Batch cultures have predominately been used for the study of physiology and gene expression in mycobacteria. This chapter describes the assembly of chemostats and the methodology that is being used for growing Mycobacterium tuberculosis in continuous culture, which provides the gr ...

The oral cavity harbors several hundred different bacterial species that colonize both hard (teeth) and soft tissues, forming complex populations known as microbial biofilms. It is widely accepted that the phenotypic characteristics of bacteria grown in biofilms are substantia ...

In their natural environments, microorganisms usually live in organized communities. Profiling analysis of microbial communities has recently assumed special relevance as it allows a thorough understanding of the diversity of the microbiota, its behavior over time, and the esta ...

Application of nucleic acid technology to the analysis of the bacterial diversity in the oral cavity in conditions of health and disease has not only confirmed the findings from early culture studies but also significantly expanded the list of oral inhabitants and candidate pathogens ass ...

Azole antifungal drugs are used widely to treat people with oral fungal infections. Unfortunately, fungi can develop resistance to these drugs. This resistance can be due to the overexpression or mutation of cytochrome P450 14α-lanosterol demethylase, also known as ERG11 or CYP51, and/or ...

For over 30 years, the chain termination method of DNA sequencing (commonly known as Sanger sequencing) has been the mainstay of any DNA sequencing project. In the past, whole-genome sequencing employing exclusively Sanger chemistry has been a labor-intensive and costly exercise and an op ...

Porphyromonas gingivalis is a Gram-negative anaerobic bacterium associated with the initiation and progression of adult periodontal disease. The pathogenicity of P. gingivalis is multifaceted and the infection process is influenced by both microbial and host factors. It is gene ...

Natural transformation is found in most groups of oral streptococci, including the mitis, the anginosus, and the mutans groups. This ability has been applied as a powerful tool to explore streptococcal gene functions and regulatory pathways, particularly in Streptococcus mutans and S ...

Most bacteria in nature exist in multispecies communities known as biofilms. In the natural habitat where resources (nutrient, space, etc.) are usually limited, individual species must compete or collaborate with other neighboring species in order to perpetuate in the multispecies c ...

Dentinal tubule invasion protects bacteria from chemo-mechanical disinfection and frequently results in root canal treatment failures. Enterococcus faecalis is a primary causative agent, particularly in persistent, asymptomatic, and chronic apical periodontitis. In o ...

Exploration of the underlying biological mechanisms of disease is useful for many purposes such as the development of novel treatment modalities in addition to informing on-going risk factor research. DNA-microarray technology is a relatively recent and novel approach to conduct ...

Both botulinum neurotoxins (BoNTs) and anthrax lethal factor, a component of anthrax toxin, exhibit zinc metalloprotease activity. The assay detailed here is capable of quantitatively detecting these proteins by measuring their enzymatic functions with high sensitivity. The d ...