Chromosome techniques: a manual

Cover......Page 1
Half Title......Page 3
Title Page......Page 5
Copyright Page......Page 6
Table of Contents......Page 7
Preface......Page 17
Introduction......Page 19
  Pre-Treabnent for Clearing the Cytoplasm and Softening the Tissue......Page 23
  Colchicine......Page 24
  Chloral Hydrate......Page 25
  Coumarin and its Derivatives......Page 26
  Monobromonaphthalene......Page 27
 Fixation......Page 30
  Non-Metallic Fixatives......Page 32
  Metallic Fixatives......Page 34
 Staining......Page 40
  Fuchsin......Page 42
  Carmine......Page 44
  Orcein......Page 46
  Crystal Violet......Page 47
  Haematoxylin......Page 48
  Giemsa (Michrome No. 144)......Page 50
  Fluorochromes......Page 51
  Temporary and Pennanent Mounts......Page 53
 A. Ordinary Light Microscopy......Page 55
 B. Phase and Interference Microscopy......Page 57
 C. Fluorescence Microscopy......Page 59
  Schedule for the Study of Chromosome Fluorescence Following Treatment with Anti-DNA Serum......Page 61
  Under Ultraviolet Light......Page 62
  Under Visible Light......Page 65
 E. Flow Cytometry......Page 68
 F. Chromosome Image Analysis......Page 69
 G. Confocal Microscopy......Page 70
Chapter 3 Study of Banding Patierns of Chromosomes......Page 71
  With Quinacrine Mustard (QM) or Quinacrine Dihydrochloride (Q)......Page 72
  With Hoechst 33258......Page 73
  Reverse Fluorescent Banding with Chromomycin and DAPI......Page 74
  Banding with Acridine Orange (AO)......Page 76
  G-Banding in Plants......Page 77
  Giemsa G-Banding in Animals......Page 78
  Feulgen-Bands......Page 81
  Giemsa C-Banding in Animals......Page 82
  Giemsa C-Banding in Plants......Page 83
 Cd-Banding......Page 85
 HKG Banding Technique for Plants......Page 86
  R-Banding in Animals......Page 87
  C. Silver Staining Methods......Page 88
 Restriction Endonuclease Banding......Page 89
  RE Banding in Human Chromosomes......Page 90
  RE Banding in Mammalian Chromosomes in Vitro......Page 91
  RE Bands in Plants (Barley)......Page 92
  In Fixed Mouse Chromosomes......Page 93
  RE-Banding and Electron Microscopy of Fixed Mouse Metaphase Chromosomes......Page 94
  Antibody Binding Methods for Human Chromosomes......Page 95
 Sister Chromatid Exchange......Page 96
  Fluorescence Plus Giemsa (FPG) Technique......Page 97
  4\'-6-Diamidino-2 Phenylindole Staining......Page 98
  Combined SCD Staining and G-Banding......Page 99
  \'Reverse\' Staining Method......Page 100
  Dot Fonnation on Chromsomes......Page 101
 A. Light Microscope Autoradiography......Page 104
  Technical Steps in the Preparation of Autoradiographs......Page 106
  Representative Schedules for Light Microscopy Autoradiography......Page 107
 C. High Resolution Autoradiography......Page 110
  Section Cutting......Page 111
  Coating of Ultra Thin Sections......Page 112
  Developing......Page 113
  Staining......Page 114
  Protocol for Spreading and Autoradiography......Page 115
 Fixation......Page 116
 Embedding......Page 118
  Schedule for Embedding with Epoxy Resin Media......Page 119
 Sectioning......Page 120
 Staining......Page 122
  EM Chromosome Analysis of Slime Mold......Page 123
  EM Analysis from Whole Mount Preparation of Protozoa......Page 124
  EM Analysis of Whole Mount Alkali Urea Treated Polytene Chromosomes......Page 125
  Study of Synaptonemal Complex......Page 126
  EM Study of Nucleosome......Page 127
  Study of Chromatin in Animals (\"Miller\" Spread)......Page 128
  Study of Chromosome Core of Trilophidia......Page 130
  Scanning Electron Microscopy......Page 131
  EM Study Following in Situ Hybridization to Metaphase Chromosomes in Animals......Page 132
  Reagents for EM......Page 135
  Principal Steps Involved in Tissue Culture......Page 137
  Representative Schedules for Plant Tissue Culture......Page 141
 B. Isolation and Culture of Protoplasts......Page 142
  Techniques for Protoplast Isolation......Page 143
 D. Fusion Between Plant Cells......Page 145
  Techniques for Cell Fusion in Plants......Page 146
  Methods for Gene Transfer......Page 147
 A. Chromosome Study from Normal Tissues......Page 152
  Study of Chromosomes from Male Germ Cells......Page 153
  Study of Chromosomes from Female Germ Cells......Page 158
  Study of Chromosomes from Somatic Cells......Page 159
  Sex Chromatin Studies......Page 169
 B. Chromosome Study from Malignant Tissues......Page 171
  Study of Cancer Tissue in Culture......Page 173
  Schedules for Preparation of Tissue......Page 174
  Schedules......Page 175
  Conclusions......Page 177
  A. Use of Sendai Virus......Page 183
  B. Virus Induced Fusion Mediated by Lectin......Page 184
  C. Use of Lipid Vesicles in Cell Fusion......Page 185
  E. Use of Polyethylene Glycol in Cell Fusion......Page 186
  F. Use of Microcells for Cell Fusion......Page 187
 Fusion Belween Plant and Animal Cells......Page 188
 Section A: Effects of Physical and Chemical Agents on Chromosomes......Page 190
  Chemical Agents......Page 191
  Schedules for Inducing Chromosome Breaks in Plants......Page 193
  Physical Agents......Page 196
  Treatment of Tissue with Both Physical and Chemical Agents......Page 201
 Section B: Methods for Monitoring Cytogenetic Effects of Environmental Toxicants......Page 204
  Some Common Test Systems Used......Page 206
  A) For Animal Cells......Page 219
  B) For Plant Tissues......Page 222
  Separation of DNA and Protein after Chromosome Isolation......Page 223
  Isolation and Separation of Heterochromatin and Euchromatin of Chromosomes......Page 225
  A) Isolation of Chromosomes in Animals......Page 227
  B) Isolation of Chromosomes in Plants......Page 230
   Mass Isolation of Chromosomes from Plant Protoplasts......Page 232
  C) Isolation of DNA from Animals......Page 233
  D) Isolation of DNA from Plants: Minipreparation......Page 239
 Micrurgical Method of Isolation of Nuclei, Chromosomes and Microdissection......Page 243
 Isolation and Culture of Nuclei in Medium under Oil......Page 244
 Microdissection of Chromosomes......Page 245
  Protocol for Microdissection......Page 246
  Microdissection of Plant Chromosomes......Page 247
Chapter 12 Identification of Chromosome Segments and DNA Sequences by In Situ Molecular Hybridization......Page 249
  In Situ Hybridization in Mammals Using Autoradiography......Page 257
  DNA In Situ Hybridization (Human)......Page 264
  In Situ Hybridization and SCE Combined Method in Human Leucocyte......Page 265
  In Situ Hybridization in Insects......Page 266
  Two-Colour Hybridization in Human Leucocyte Culture......Page 267
  Multiple Banding and In Situ Hybridization in Mammal......Page 270
  MICRO-FISH—Rapid Regeneration of Probes by Chromosome Microdissection......Page 271
  In Situ Hybridization Using Radioactive Probe in Plant Pachytene......Page 273
  Genomic In Situ Hybridization with Non-Radioactive Probe (GISH) for Plants......Page 274
  In Situ Hybridization with Automated Chromosome Denaturation in Plants......Page 278
  Mapping of Low Copy DNA Sequence in Plants......Page 280
  In Situ Hybridization (Plants) with Biotin-Labelled Probe......Page 281
  Two Colour Mapping of Plant DNA Sequences......Page 282
  In Situ Hybridization with Immunofluorescent Antibodies......Page 283
  Monoclonal Antibody Against Kinetochore Component — Schedule for Preparation in CHO Cells......Page 284
  Indirect Immunofluorescence in Mammalian Sperm......Page 286
  Modified Banding of Human Chromosomes for Electron Microscopy......Page 287
  Antibody Labelling and Flow Cytometry in CHO Cell Line......Page 288
  Antigen Reaction with Human Autoantibody in Diptera......Page 289
  Anti-Topoisomerase II in Recognising Meiotic Chromosome Core in Bird......Page 290
  BrdU Antibody Technique (BAT)......Page 291
  Products Available for In Situ Hybridization......Page 292
  Kinetochore Visualization with Crest Serum in Plants......Page 293
  Methylation of DNA......Page 294
 Section A: Restricfion Fragment Length Polymorphism......Page 296
  Preparation and Examination of Agarose Gels......Page 300
  Separation of Restriction Fragments of Mammalian Genomic DNA by Agarose Gel Electrophoresis......Page 302
  Hybridization of Radiolabeled Probes to Nucleic Acids Immobilized on Nitrocellulose Filters or Nylon Membranes......Page 305
 Section B: The DNA Fingerprint......Page 307
 Section C: Polymerase Chain Reaction......Page 310
 Section D: Chromosome Walking......Page 315
 Section E: Foot Print......Page 320
  Schedules for the Study of Mitotic Chromosomes......Page 321
  Schedules for the Study of Meiotic Chromosomes......Page 331
  1. Chromosome Segments......Page 335
  2. Special Chromosomes......Page 337
  3. Special Groups......Page 343
  4. Components-Nucleic Acids......Page 349
References......Page 357
Index......Page 389