Analyzing Chromosomes - B. Czepulkowski (BIOS, 2001) WW

221 Pages·2001·14.04 MB·English

Analyzing
Chromosomes

B. Czepulkowski

The

BASICS rom bactround to bane
Analyzing
Chromosomes
The

asics

Series Advisors:

Rob Beynon UMIST, Manchester, UK
Chris Howe Department of Biochemistry, University of Cambridge, Cambridge, UK

Monoclonal Antibodies
PCR

Separating Cells
Analyzing Chromosomes

Forthcoming titles
Biological Centrifugation

Gene Mapping
Reconstructing Molecular Evolutionary Trees
Analyzing
Chromosomes

Barbara Czepulkowski

Leukaemia Sciences Laboratories
The Rayne Institute, London, UK
© BIOS Scientific Publishers Limited, 2001
First published 2001

All rights reserved. No part of this book may be reproduced or transmitted, in any form or by any means,
without permission.

A CIP catalogue record for this book is available from the British Library.

ISBN 1 85996 188 6

BIOS Scientific Publishers Ltd

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Published in the United States of America, its dependent territories and Canada by Springer-Verlag New
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Production Editor: Andrea Bosher
Typeset by Creative Associates, Oxford, UK
Printed by The Cromwell Press, Trowbridge, UK
Abbreviations
Contributors
Introduction

Chapter |

Contents

Introducing the cell

Barbara Czepulkowski

A a

Introduction

The cytoplasm

The nucleus

DNA replication

Mutation and DNA repair

Chromosome organization and structure
Chromosome function

Chapter 2 Culture of human cells for
chromosomal analysis

Karen Saunders and Barbara Czepulkowski

SA pe wy

Introduction

Culture of peripheral blood samples

Culture of cells from amniotic fluid

Culture of chorionic villus samples

Solid tissue culture

Culture of bone marrow and leukemic blood

Culture of samples from chronic lymphoid leukemia and

the lymphomas
8. Slide preparation

Protocol 2.1:
Protocol 2.2:
Protocol 2.3:
Protocol 2.4:
Protocol 2.5:
Protocol 2.6:
Protocol 2.7:

Protocol 2.8:

Protocol 2.9:

Protocol 2.10:

Setting up cell cultures and harvesting
Amniotic fluid culture

Harvesting amniocytes

Thymidine blocking

Addition of BrdU with colcemid

Sorting of the chorionic villus sample

Direct culture of chorionic villus samples
Long-term culture of chorionic villus samples
Setting up bone marrow and leukemic blood
Synchronization of bone marrow cultures using a
BrdU/FdU cocktail mixture

xiii
vi Analyzing chromosomes

Protocol 2.11: Harvest of bone marrow and blood cultures 43
Protocol 2.12: Culture of samples from CLL blood and marrow 45
Protocol 2.13: Preparation and culture of samples from lymphomas 46
Protocol 2.14: Slide-making 47

Chapter 3 Staining and banding of
chromosome slides 49

Barbara Czepulkowski

1. Introduction 49
2. Banding techniques and their applications 50
Protocol 3.1: Solid staining 53
Protocol 3.2: Preparation of Leishman’s stain for G-banding 55
Protocol 3.3: G-banding 56
Protocol 3.4: Q-banding 59
C-banding 61
R-banding 63
Protocol 3.7: Nuclear organizer region (NOR) staining 65
Protocol 3.8: DAPI/distamycin A staining 67
Chapter 4 Types of abnormalities observed
in chromosomes 69

Barbara Czepulkowski

1. Introduction 69
2. Types of chromosomal changes 69
3. Nomenclature for human chromosomes 78
Chapter 5 Approaching analysis a
Barbara Czepulkowski
1. Introduction a1
2. Guidelines for the analysis of chromosomes from

constitutional studies 96
3. Guidelines for the analysis of chromosomes from

malignant tissues 103
4. Clonal abnormality 106

Chapter 6 Constitutional chromosome
abnormalities 109

Barbara Czepulkowski and Karen Saunders

1. Introduction 109
2. Trisomy 21 - Down syndrome mw
3. Trisomy 18 - Edward syndrome 12
4, Trisomy 13 - Patau syndrome 13
5, 45,X - Turner syndrome 13
6. 47,XXY - Klinefelter syndrome 14
7. 46,XX males 5
Contents vii

8. 47,XYY us

9. 47,XXX 116
10. Triploidy 116
11. Microdeletion syndromes and chromosomal duplications 116
12. Marker chromosomes 17
13. Ring chromosomes 120
14. Fragile X syndrome 121

Chapter 7 Acquired chromosome abnormalities

observed in malignancy 125
Barbara Czepulkowski
1. Introduction 125
2. Classification and clinical symptoms of myeloid malignancies 126
3. Chromosomal abnormalities observed in acute myeloid leukemia 130
4. Chromosome changes in myelodysplastic syndromes 136
5. Chromosome changes in myeloproliferative disorders 138
6. Acute lymphoblastic leukemia 141
7. Chronic lymphoproliferative disorders and lymphomas 149
8. Solid tumors 155
9. Summary 156
Chapter 8 New complementary techniques 159
Karen Saunders and David Jones
1. Introduction 159
2. Fluorescent in situ hybridization 159
3. Twenty-four color painting 167
4. Color banding 168
5. Primed in situ hybridization 169
6. Fiber FISH 169
7. Comparative genomic hybridization 169
8. The present and future of cytogenetics 171
Protocol 8.1: Processing of uncultured blood and bone marrow

preparations for interphase FISH 173
Protocol 8.2: Processing of uncultured amniocytes for

interphase FISH 174
Protocol 8.3: Pre-treatment of slides for uncultured samples

(or very cytoplasmic samples) 176
Protocol 8.4: Standard protocol for in situ hybridization 178
Appendix | 183
Appendix I 191
Index 199

The color section can be found between pages 18 and 19