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Provides detailed protocols for studying the molecular biology of the pathogen M. tuberculosis and its interactions with host cells. Both traditional and novel methodologies are reviewed, with chapters covering everything from safety issues and the storage of strains to the application of the latest functional genomics technologies. These molecular techniques are suitable for research in genetics, cell biology, biochemistry, microbiology, epidemiology, and diagnostics. Techniques include methods for the basic safety and culture of M. tuberculosis, fractionation of the bacterium, analysis of gene expression, and cytological analysis. Parish is affiliated with St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary and Westfield College, UK. Stoker is affiliated with the London School of Hygiene and Tropical Medicine, UK.Book News, Inc.

Book Info

Royal London School of Medicine and Dentistry, UK. Provides detailed protocols for studying the molecular biology of the pathogen Mycobacterium tuberculosis, and its interactions with host cells. Topics range from storage of strains and safety issues to the application of functional genomics technologies.

Preface
Contributors
1 Plasmid Vectors 1
2 Isolation of DNA from Mycobacterium tuberculosis 19
3 Extraction of RNA from Intracellular Mycobacterium tuberculosis: Methods, Considerations, and Applications 31
4 Transposon Mutagenesis in Mycobacteria Using Conditionally Replicating Mycobacteriophages 43
5 Gene Replacement and Transposon Delivery Using the Negative Selection Marker sacB 59
6 Gene Replacement Using Pretreated DNA 77
7 Gene Replacement in Mycobacterium tuberculosis and Mycobacterium bovis BCG Using rpsL[superscript +] as a Dominant Negative Selectable Marker 93
8 Transcription Start-Site Mapping 105
9 Fluorescence and Brightfield Cytology of Live M. tuberculosis Cells 125
10 Phage Replication Technology for Diagnosis and Drug Susceptibility Testing 145
11 Detection of Mutations of Mycobacterium tuberculosis by a Dot Blot Hybridization Strategy 153
12 Restriction Fragment Length Polymorphism Typing of Mycobacteria 165
13 Preparation of Culture Filtrate Proteins from Mycobacterium tuberculosis 205
14 Analysis of the Capsule of Mycobacterium tuberculosis 217
15 Analysis of the Lipids of Mycobacterium tuberculosis 229
16 In Vitro Model of Hypoxically Induced Nonreplicating Persistence of Mycobacterium tuberculosis 247
17 Macrophage Virulence Assays 271
18 Analysis of Mycobacterium-Infected Macrophages by Immunoelectron Microscopy and Cell Fractionation 281
19 Real-Time PCR Using Molecular Beacons: A New Tool to Identify Point Mutations and to Analyze Gene Expression in Mycobacterium tuberculosis 295
20 Electronic Access to Mycobacterium tuberculosis Sequence Data 311
21 Proteomics 315
22 Functional Genomics of Mycobacterium tuberculosis Using DNA Microarrays 335
23 Storage of Mycobacterial Strains 359
24 Safety in the Laboratory 367
Index 385