Chemical Mutagens: Principles and Methods for Their Detection. Vol. 2

The best protection against environmental mutagens is to identify them before they ever come into general use. But it is always possible that some substance will escape detection and affect a large number of persons without this being realized until later generations. This article considers ways in which such a genetic emergency might be promptly detected. A mutation-detecting system should be relevant in that it tests for effects that are as closely related as possible to those that are feared. It should be sensitive enough to detect a moderate increase in mutation rate, able to discover the increase promptly before more damage is done, responsive to various kinds of mutational events, and designed in such a way as to maxi­ mize the probability that the Gause of an increase can be found. Methods based on germinal mutation necessarily involve enormous numbers of persons and tests. On the other hand, with somatic mutations the individual cell becomes the unit of measurement rather than the in­ dividual person. For this reason, I think that somatic tests are preferable to germinal tests, despite the fact that it is germinal mutations which are feared.

of Volume 2.- 11 Measurement of Recessive Lethal Damage Over the Entire Genome and at Two Specific Loci in the ad-3 Region of a Two-Component Heterokaryon of Neurospora Crassa.- I. Introduction.- A. Use of a Two-Component Heterokaryon to Measure the Genetic Effects of Mutagenic Treatment.- B. The Spectrum of Recessive Lethal Mutations Detectable with a Two-Component Heterokaryon.- C. Characteristics of Strain 12-A Two-Component Heterokary on.- D. Assaying for Mutagenicity of Chemicals in Screening Programs.- II. Measurement of the Genetic Effects of Mutagenic Treatment.- A. General Methods of Treatment.- B. Tests for Mutagenicity on Growing Cultures.- C. Tests for Mutagenicity on Nongrowing Conidia.- D. Treatment with Mutagens.- E. Assaying for the Genetic Effects of Mutagenic Treatment.- F. Evaluation of Genetic Assays.- III. Characterization of Presumptive ad-3 Mutants.- A. Establishing a Silica Gel Stock Culture of Each Adenine-Requiring Strain.- B. Genetic Tests.- IV. Media and Chemical Solutions.- A. Media.- B. Chemical Solutions.- V. References.- 12 Aspergillus.- I. Introduction.- II. Life Cycle and Genetic Analysis.- A. The Vegetative Cycle.- B. The Sexual Cycle.- C. The Parasexual Cycle.- III. Mutation.- A. Gene Mutations.- B. Mutations of Quantity and Arrangement.- IV. Conclusions.- V. References.- 13 Higher Plants.- I. Introduction.- II. Test Systems.- A. Barley.- B. Pea.- C. Wheat and Other Polyploids.- D. Arabidopsis thaliana.- III. Compounds Tested for Mutagenic Activity by Treatment of Seeds.- A. Classification of Mutagens.- B. Quantitative Aspects.- C. Alkylating Agents.- D. Potential Unspecific Mutagens Active by Mechanisms Other Than Alkylation.- E. Agents Causing Specific Changes in DNA Bases.- IV. Special Techniques That Have a Higher Resolving Power.- A. Pollen Characters.- B. Somatic Mutant Sectors.- V. General Evaluation.- VI. Acknowledgments.- VII. References.- 14 Procedures for Culturing Diploid Cells and Preparation of Meiotic Chromosomes from Dwarf Species of Hamsters.- I. Introduction.- II. Background Information.- A. Brief Historical Account.- B. Breeding and Management.- C. General Characteristics of Animals and Cultured Cells.- D. Cytological Preparations.- E. Spontaneous Tumors.- III. Handling the Diploid Cell.- A. Cloning Operations.- B. Difficulties When Employing Single Type Serum Supplemented Media.- C. Cell Culture Media for Routine Propagation of Different Diploid Cell Types.- D. Combined Sera Supplements.- E. Phenotypic Features of Diploid Cells.- F. Initiation of Cell Cultures from Organs and Solid (Primary) Tumors.- G. Derivation of Diploid Cell Lines from Peritoneal Exudate.- H. Roller-Flask Cultures of Diploid Cell Lines.- IV. Spermatogonial and Meiotic Chromosome Preparations.- A. Partial Orchidectomy.- B. Hypotonic Treatment and Fixation.- C. Aceto-Carmine Squash Preparations.- D. Propiono-Carmine Squash Preparation.- E. Air-Dried Preparations.- F. Construction of a Low-Cost Centrifuge Tube “Flicker”.- G. Preparation of Lactic-Acetic-Orcein.- H. Permanent Slides.- V. Bone Marrow Biopsy Procedure.- VI. Discussion.- VII. Acknowledgments.- VIII. References.- 15 Induction and Analysis of Gene Mutations in Mammalian Cells in Culture.- I. Introduction.- II. The Cell Material.- A. Species Origin.- B. Karyotypic Stability.- C. Plating or Cloning Efficiency.- D. Cold Storage of Cells.- III. Utilization of In Vivo Markers.- A. Morphological Markers.- B. Biochemical Markers.- C. Serological Markers.- D. Radiation-Sensitive Mutants.- IV. Detection of Recessive Mutations in Cell Cultures.- A. Heterozygosity of Autosomal Recessive Genes.- B. Aneuploidy for Autosomes.- C. Natural Monosomy of X Chromosomes in Normal Males or in XO Females.- D. Functional Monosomy of X Chromosomes in Normal Females.- E. Independent Mutation at Both Alleles in Homozygous Dominant Genes.- V. Selective Techniques for New Mutations in Cell Culture.- A. Mass Selection Method.- B. Lethal-Growth Method.- C. “Thymineless Death” Method.- D. Replica-Plating Method.- VI. Characterization of Newly Isolated Variants.- VII. Procedure for Mutation Induction.- A. Choice of the Test System.- B. Forward and Back Mutations.- C. Determination of Cytotoxicity.- D. Elimination of Background Mutations.- E. Treatment with Mutagen.- F. Inoculation of Cells.- G. Addition of Selective Agent.- H. Isolation and Testing of Mutant Colonies.- I. Fixation and Staining of Colonies.- J. Mutation Rate and Mutation Frequency.- VIII. Concluding Remarks.- IX. Acknowledgments.- X. References.- 16 Inducing Mutations with Chemicals in Habrobracon.- I. Introduction.- II. Maintenance of Habrobracon and Ephestia.- A. Habrobracon Culture.- B. Ephestia Culture.- III. Scheme for Mutational Analysis.- A. General Stock.- B. Pretest.- C. Collection of Virgins and Males for Experiment.- D. Treatments.- E. Counts.- F. Test of F1 Females.- G. Mating and Further Testing of Females with Genetic Alterations.- H. Special Mutant Analysis.- I. Homozygosity of Mutants.- J. Further Testing.- K. Data Processing.- L. Catalog of Mutants.- IV. Application of Mutagens.- A. Aerosols.- B. Feeding.- C. Topical Application.- D. Microinjection.- V. Conclusions.- VI. References.- 17 The Detection of Mutations in Drosophila melanogaster.- I. Introduction.- II. Advantages of Drosophila as a Test Organism.- III. Limitations of Drosophila as a Test Organism.- IV. Lethal Tests.- A. Sex-Linked Recessive Lethals.- B. Autosomal Recessive Lethal Test.- V. Tests for Recessive Visible Mutants.- VI. Tests for Chromosomal Rearrangements.- A. Genetic Test for Reciprocal Translocation.- B. Position-Effect Tests for Chromosomal Rearrangement.- C. Cytological Test for Chromosomal Rearrangement.- VII. Tests for Loss of X or Y Chromosomes.- VIII. Tests for Dominant Lethals.- IX. Tests for Half-Translocations.- X. Staging of Germ Cells.- XI. Techniques for Collecting Flies of a Desired Sex.- XII. Review of Literature.- XIII. Summary.- XIV. References.- 18 Root Tips for Studying the Effects of Chemicals on Chromosomes.- I. The Material.- A. Root Tips as Experimental Material.- B. The Horse Bean, Vicia faba.- C. The Common Onion, Allium cepa.- D. The Tree Onion, Allium proliferum.- II. Treatment of Root Tips with Chemicals.- III. Fixation and Staining of Root Tips.- IV. Scoring of Slides and Types of Aberration.- V. Comparison Between the Effects of Chemicals on Chromosomes in Root-Tip Cells and in Cultured Animal Cells.- A. Are Results Obtained in Root Tips Representative for Other Materials?.- B. Tepa and Related Compounds.- C. Nitrilotriacetic Acid, NTA.- D. Caffeine.- E. Concluding Remarks.- VI. Acknowledgments.- VII. References.- VIII. Suggested Reading.- 19 Cytogenetic Studies in Animals.- I. Introduction.- II. Experimental Design.- A. Controls.- B. Replication.- C. Observer Bias.- D. Standardizing Scoring Methods.- E. Statistical Evaluation.- F. Types of Damage.- III. Classification of Chromosomal Aberrations.- IV. Localization of Chromosomal Aberrations.- V. Timing of Chromosomal Damage.- VI. Meiotic Studies.- VII. In Vivo and in Vitro Studies.- VIII. Specific Techniques.- A. Lymphocyte Culture.- B. Bone Marrow (Direct).- C. Fibroblast Culture.- D. Amniotic Fluid Cell Culture.- E. Meiotic Preparations.- IX. Summary.- X. References.- 20 Specific Locus Mutation in Mice.- I. Introduction.- II. The Method, Its Advantages and Disadvantages.- III. Results Obtained.- IV. Conclusions.- V. References.- 21 Dominant Lethal Mutations in Mammals.- I. Introduction.- II. The Dominant Lethal Syndrome.- A. Critical Stages of Pregnancy.- B. Modes of Egg or Fetal Death.- III. The Estimation of Dominant Lethals.- A. Mid-Term Litters.- B. Full-Term Litters.- C. Recommended Protocols.- IV. The Genotypes of Dominant Lethals.- A. Chromosome Loss.- B. Monosomy and Trisomy.- Y. Dominant Lethals in Male Germ Cells.- A. Sperm Maturation.- B. Differential Sensitivity.- VI. Dominant Lethals in Female Germ Cells.- VII. General Validity of the Dominant Lethal Test.- VIII. Synergistic Effects.- IX. Review of Chemicals Tested as Dominant Lethal Mutagens in Mammals.- X. Integration of the Dominant Lethal Assay and Other Mutagenicity Tests into General Toxicological Practice.- XI. References.- 22 The Host-Mediated Assay, a Practical Procedure for Evaluating Potential Mutagenic Agents in Mammals.- I. Introduction.- II. Materials and Methods.- A. Strains.- B. Host-Mediated Assay.- III. Results.- A. Salmonella.- B. Neurospora.- C. Comparative Mutagenicity.- IV. Conclusion.- V. References.- 23 Human Population Monitoring.- I. Introduction.- II. A Classification of Mutational Effects.- A. Cytogenetic Changes.- B. Dominant Mutations.- C. X-Linked Recessive Mutations.- D. Autosomal Recessive Mutations.- E. Mutants with Minor Effects.- III. Criteria for a Mutation-Monitoring System.- A. Is the System Relevant?.- B. How Quickly Will a Mutation Increase Be Detected?.- C. Can the System Detect a Small Increase in the Mutation Rate?.- D. Can Many Kinds of Mutational Events Be Detected?.- E. Does the System Offer a High Probability of Identifying the Cause of the Mutation Increase?.- F. Is the System Available Now?.- IV. Some Ways of Amplifying the Mutation-Detecting Power of Monitoring Systems.- A. Somatic Cytogenetic Studies.- B. Somatic Mutation-Detection Systems.- C. Indirect Monitoring by Testing for Mutagens in Human Blood.- V. Monitoring for Germinal Mutations.- A. Monitoring for Dominant Mutant Phenotypes.- B. Biochemical Monitoring.- VI. Summary.- VII. References.- Conclusion.- Author Index.