Technologies for Detection of DNA Damage and Mutations

Man-made carcinogens, natural genotoxic agents in the environment, as well as ionizing and ultraviolet radiation can damage DNA and are a constant threat to genome integrity. Throughout the evolution oflife, complex DNA repair systems have developed in all living organisms to cope with this damage. Unrepaired DNA lesions can promote genetic alterations (mutations) that may be linked to an altered phenotype, and, if growth-controlling genes are involved, these mutations can lead to cell transformation and the development of malignant tumors. Proto­ oncogenes and tumor suppressor genes may be critical targets for DNA damaging agents. In a number of animal model systems, correlations between exposure to a carcinogen, tumor develop­ ment, and genetic changes in tumor DNA have been established. To understand mutagenesis processes in more detail at the molecular level, we need to know the type and frequency of DNA adducts within cells, their distribution along genes and specific DNA sequences, as well as the rates at which they are repaired. We also need to know what types of mutations are produced and which gene positions are most prone to mutagenesis. This book provides a collection of techniques that are useful in mutagenesis research. The book is divided into three parts. In Part I, methods for DNA damage and repair analysis are provided.

Table of Contents:
I. Technologies for Detection of DNA Damage.- 1. Microgel Electrophoresis of DNA from Individual Cells: Principles and Methodology.- 2. The Cytokinesis-Block Micronucleus Technique.- 3. Agarose Gel Electrophoresis for DNA Damage Analysis.- 4. 32P-Postlabeling for Detection of DNA Adducts.- 5. A Postlabeling Assay for Oxidative Damage.- 6. Radioimmunoassay of DNA Damaged by Ultraviolet Light.- 7. Monoclonal Antibody-Based Quantification and Repair Analysis of Specific Alkylation Products in DNA.- 8. Detection of Oxidative DNA Base Damages: Immunochemical and Electrochemical Approaches.- 9. Strategies for Measuring Damage and Repair in Gene-Sized Specific DNA Sequences.- 10. Methods to Measure the Repair of Genes.- 11. Mapping and Quantification of Bulky Chemical-Induced DNA Damage Using UvrABC Nucleases.- 12. The Use of DNA Glycosylases to Detect DNA Damage.- 13. PCR-Based Assays for the Detection and Quantitation of DNA Damage and Repair.- 14. DNA Damage Analysis Using an Automated DNA Sequencer.- 15. Ligation-Mediated PCR for Analysis of UV Damage.- 16. Ligation-Mediated PCR for Analysis of Oxidative DNA Damage.- 17. Single-Strand Ligation PCR for Detection of DNA Adducts.- II. Technologies for Detection of Mutations.- 18. Heteroduplex Analysis.- 19. Mutation Analysis by Denaturing Gradient Gel Electrophoresis (DGGE).- 20. Constant Denaturant Gel Electrophoresis (CDGE) in Mutation Screening.- 21. Single-Strand Conformation Polymorphism Analysis.- 22. Two-Dimensional Gene Scanning.- 23. Ligase Chain Reaction for the Detection of Specific DNA Sequences and Point Mutations.- 24. The Protein Truncation Test (PTT) for Rapid Detection of Translation-Terminating Mutations.- 25. Single Nucleotide Primer Extension for Analysis of Sequence Variants.- 26. Sequencing of PCR Products.- III. Mammalian Systems for Mutation Analysis.- 27. Detection and Characterization of Mutations in Mammalian Cells with the pSP189 Shuttle Vector System.- 28. The HPRT Gene as a Model System for Mutation Analysis.- 29. Bacteriophage Lambda and Plasmid lacZ Transgenic Mice for Studying Mutations in Vivo.- 30. The Use of lacI Transgenic Mice in Genetic Toxicology.- 31. Genotypic Mutation Assay (RFLP/PCR).