Strategies needed to isolate mutant cell lines and to use the mutants to clone and map the genes are presented in detail. This is the first book in which such methodology and molecular biology are combined.The preferred cell lines for genetic analysis are presented in the first section, followed by special techniques for isolating a wide range of mutants. The remainder of the book is devoted to genetic mapping and cloning of mutant genes. The final section presents special techniques in gene regulation.
Author(s): Michael M. Gottesman
Series: Methods in Enzymology 151
Publisher: Academic Press
Year: 1988
Language: English
Commentary: index pages missing
Pages: 624
Table of Contents......Page 6
Contributors to Volume 151......Page 10
Menashe Marcus......Page 14
Preface......Page 16
VOLUMES IN SERIES......Page 18
Section I Cell Lines Useful for Genetic Analysis......Page 32
Chinese Hamster Ovary Cells......Page 34
Cultured 549 Mouse T Lymphoma Cells......Page 40
Liver Cell Lines......Page 50
HeLa Cell Lines......Page 69
Somatic Cell Genetic Analysis of Myelomas and
Hybridomas......Page 81
Mouse Teratocarcinoma Cells......Page 97
Section II Special Techniques for Mutant Selection......Page 114
Amplification of Genes in Somatic Mammalian
Cells......Page 116
Replica Plating and Delection of Antigen Variants......Page 135
Drug-Resistant Mutants: Selection and and Dominance Analysis......Page 144
Suicide Selection of Mammalian Cell Mutants......Page 152
Use of Tritium-Labeled Precursors to Select
Mutants......Page 162
Construction of Immunotoxins Using Pseudomonas Exotoxin A......Page 170
Isolation of Temperature-Sensitive Mutants......Page 176
Fluorescence-Activated Cell Sorting of Hybrid and Transfected Cells......Page 181
Section III Genetic Mapping and Analysis......Page 198
Isozyme Identification of Chromosomes inInterspecific Somatic Cell Hybrids......Page 200
Principles of Electrofusion and Electropermeabilizalion......Page 225
Cell Enucleation. Cybrids. Reconstituted Cells.
and Nuclear Hybrids......Page 256
Toxin-Antiloxin Selection for Isolating Helerobryonsand Cell Hybrids......Page 272
Chromosome Sorting by Flow Cytometry......Page 287
Methods for Chromosome Banding of Human and
Experimental Tumors in Vitro......Page 302
In Silll Hybridization of Metaphase and Prometaphase Chromosomes......Page 314
Gene Mapping with Sorted Chromosomes......Page 327
Transfer and Selective Retention of Single Specific
Human Chromosomes via Microcell·Mediated
Chromosome Transfer......Page 348
Section IV Isolation and Detection of Mutant Genes......Page 362
Strategies for Isolation of Mutant Genes......Page 364
Use of in-Gel DNA Renaturation for Detection and Cloning of Amplified Genes......Page 367
Detection and Characterization or Specific mRNA by Microinjection and Complementation of Mutant Cells......Page 406
Use or the CAT Reporter Gene ror Optimization of Gene Transfer into Eukaryotic Cells......Page 417
Cosmid Vector Systems for Genomic DNA Cloning......Page 432
Preparation of a Genomic Cosmid Library......Page 440
Colony Screening orGcnomic Cosmid Libraries......Page 451
Rescue of Genes Transferred from Cosmid Libraries......Page 461
Phage-Mediated Transduction of cDNA Expression Libraries into Mammalian Cells......Page 469
Sib Selection......Page 480
Repetitive Ooning or Mutant Genes Using Locus-Specific Sticky Ends......Page 484
Strategies ror Mapping and Cloning Macroregions of Mammalian Genomes......Page 496
Insertional Mutagenesis to Isolate Genes......Page 524
Microcloning of Mammalian Metaphase Chromosomes......Page 538
Section V Gene Regulation in Tissue Culture......Page 552
Antisense RNA Inhibition or Endogenous Genes......Page 554
In Situ Detection or Transcription in Transrected Cells Using Biotin·Labeled Molecular Probes......Page 565
RNA Delection and Localization in Cells and Tissue Sections by in Situ Hybridization of 35S-Labeled RNA Probes......Page 574
Electrophoretic Assay for DNA-Binding Proteins......Page 586
Author Index......Page 602
