DNA Fingerprinting: An Introduction PDF
Preview DNA Fingerprinting: An Introduction
DNA Fingerprinting An Introduction Breakthroughs In Molecular Biology DNA Fingerprinting is the second volume to appear in this exciting new series of high quality, affordable books on the fields of molecular biology and immunology. This series is dedicated to the rapid publication of the latest breakthroughs and cutting edge technologies as well as syntheses of major advances within molecular biology. Other volumes in the series include: PCR Technology: Principles and Applications for DNA Amplification edited by H. Erlich Lymphokines: The Molecular Biology of Regulators of Immune and Inflammatory Responses by K.-I. Arai and J. de Vries Homologous Recombination and Gene Targeting by J. Sedivy and A. Joyner Signal Transduction in Biological Systems by Y. Kaziro and K.-I. Arai YAC Libraries: Construction and Use edited by B. Brownstein DNA Fingerprinting An Introduction Lorne T. Kirby M Palg rave Macmillan ©Stockton Press, 1990 All rights reserved. No part of this publication may be reproduced, or transmitted, in any form or by any means, without permission. Published in the United States and Canada by Stockton Press 15 East 26th Street, New York, N.Y. 10010 Library of Congress Cataloglng·ln·Publicatlon Data DNA fingerprinting: an introduction/Lorne T. Kirby. p. cm.-(Breakthroughs in molecular biology) Includes bibliographical references. ISBN 978·0·935859·94·2 (soft) 1. DNA fingerprints. I. Kirby, Lorne T., 1938- . II. Series. [DNLM: 1. DNA-genetics. 2. Forensic Medicine. au 58 062825] RA1057.55.D635 1990 614'.1-dc20 DNLMlDLC for Library of Congress 90-9527 CIP ISBN 978-0-935859-94-2 Published in the United Kingdom by MACMILLAN PUBLISHERS LTD (Journals Division), 1990 British Library Cataloguing In Publication Data DNA Fingerprinting 1. Forensic medicine. Role of organisms I. Kirby, Lorne T. 1938- 614.1 IISSBBNN 9 97788-0-0-3-33333-5-45042042-44- 4 I S B INS B9N78 9-17-83-419-3-14290-4102-064 (0e-B6o (oekB) ook) 0D0O1I1 100.1.010070/79/7987-81--13-4394-91-21024004-06- 6 987654321 To William and Mary Kirby, Sarah-Dean, Christopher, and Marcia Louise CONTENTS 1. INTRODUCTION ................................................................................ 1 DEFINITION ........................................................................................ 1 mSTORY ............................................................................................. 2 A CASE FOR DNA .............................................................................. 3 APPLICATIONS .................................................................................. 3 REFERENCES ...................................................................................... 5 2. GENETIC PRINCIPLES ..................................................................... 7 DNA STRUCTURE .............................................................................. 8 Nuclear DNA .................................................................................... 8 Organelle DNA ............................................................................... 10 DNA FUNCTION ............................................................................... 11 REPRODUCTION .............................................................................. 13 LINKAGE ........................................................................................... 19 PEDIGREE ANALYSIS ..................................................................... 20 MUTATION ....................................................................................... 20 RESTRICTION ENDONUCLEASES ................................................ 23 CONCEPT OF POLyMORPHISM .................................................... 24 RECOMBINANT DNA ...................................................................... 26 BASE SEQUENCE DETERMINATION ........................................... 29 OLIGONUCLEOTIDE SYNTHESIS ................................................ 30 REPETITIVE SEQUENCE ORGANIZATION ................................. 31 NOMENCLATURE OF ARBITRARY DNA SEGMENTS .............. 33 REFERENCES .................................................................................... 33 vii viii Contents 3. LABORATORY ORGANIZATION ................................................. 35 LABORATORY SECTIONS ............................................................. 36 Office .............................................................................................. 36 Specimen Documentation .............................................................. 36 Specimen Processing ...................................................................... 36 General Preparation ........................................................................ 39 Vector Handling ............................................................................. 39 Amplification by PCR .................................................................... 39 Hybridization and Radioisotope Handling ..................................... 39 Darkroom ........................................................................................ 45 Result Interpretation ....................................................................... 45 Tissue Culture ................................................................................. 45 Wash-up Area ................................................................................. 45 EQUIPMENT SUMMARY ................................................................ 46 OPERATION ...................................................................................... 47 Personnel ........................................................................................ 47 WorkFlow ...................................................................................... 48 Safety .............................................................................................. 49 REFERENCES .................................................................................... 50 4. SPECIMENS ........................................................................................ 51 COLLECTION .................................................................................... 52 STORAGE AND TRANSPORT ........................................................ 55 DNA EXTRACTION .......................................................................... 55 Principles ........................................................................................ 55 Automated System ......................................................................... 56 Manual System Organic Solvent Extraction .................................. 56 Whole blood (Method 1) • Whole blood (Method 2) • Tissue culture, amniotic fluid, and buccal cells· Hair roots, biopsy, and autopsy tissues· Fixed tissues· Stains· Sperm and female-cell mixtures· Plant tissues Manual System Nonorganic Solvent Extraction ............................ 65 Whole blood· Stains • Sperm and female-cell mixtures DIALySIS ........................................................................................... 66 RNAASE TREATMENT ...................................................................... 67 QUANTITATION ............................................................................... 67 DNA CONCENTRATION ................................................................. 68 QUALITY DETERMINATION ......................................................... 69 ix EFFECTS OF ENVIRONMENTAL FACTORS INCLUDING CONTAMINATING DNA ON DNA PROFILES ...... 69 ANALYSIS REAGENTS ................................................................... 70 REFERENCES .................................................................................... 73 5. DNA AMPLIFICATION .................................................................... 75 POLYMERASE CHAIN REACTION ............................................... 76 Amplified Fragment Length Polymorphisms (AMP-FLPs) ........... 77 Advantages of PCR ........................................................................ 78 Pitfalls in PCR ................................................................................ 78 A PCR Protocol .............................................................................. 78 PROBE AMPLIFICATION ................................................................ 79 TISSUE CULTURE ............................................................................ 82 REFERENCES .................................................................................... 89 6. ANALYSIS TECHNIQUES ............................................................... 91 RESTRICTION ENZYME CLEAVAGE ........................................... 94 GEL-BLOT ORGANIZATION .......................................................... 95 Forensic .......................................................................................... 95 Parentage ........................................................................................ 95 Size Markers ................................................................................... 96 ELECTROPHORESIS ........................................................................ 96 Equipment ...................................................................................... 97 Reagents ......................................................................................... 97 Procedure ........................................................................................ 98 PHOTOGRAPHY ............................................................................... 99 DNA RECOVERY ............................................................................ 100 MEMBRANE TRANSFER .............................................................. 100 Southern Blots .............................................................................. 100 Dot-(Slot-) Blots ........................................................................... 104 RADIOISOTOPE PROBE LABELING ........................................... 104 NONRADIOISOTOPE PROBE LABELING .................................. 109 HYBRIDIZATION ........................................................................... 110 Principles ...................................................................................... 111 Procedures .................................................................................... 112 PCR Products ............................................................................... 115 PRINT DETECTION ........................................................................ 115 x Contents BLOT STRIPPING AND REPROBING .......................................... 116 DATA PROCESSING ...................................................................... 116 Allele Fragment Size Determination ............................................ 116 Digital Analysis ............................................................................ 118 The FBI System ............................................................................ 118 Interpretation of Profile Matches ................................................. 119 PROFILE REP<)RT .......................................................................... 125 DNA PROFILE DATA BASES ....................................................... 126 P<)TENTIAL PITFALLS .................................................................. 127 TROUBLESHOOTING GUIDE ...................................................... 130 ANALYSIS COST ............................................................................ 130 ANALYSIS REAGENTS ................................................................. 130 REFERENCES .................................................................................. 131 7. PROBES, ALLELE MUTATIONS, AND RESTRICTION ENZYMES ............................................................ 135 PROBES ............................................................................................ 136 Polymorphism .............................................................................. 136 Tandem Repeat Single-Locus vs. Multilocus Probes ................... 13 7 Oligonucleotide Probes ................................................................ 139 Amplification and Stability .......................................................... 139 Availability ................................................................................... 141 Probes Used in Forensic Analysis ................................................ 141 MUTATIONS ................................................................................... 142 Allele Formation .......................................................................... 142 Genotype Analysis ....................................................................... 142 RESTRICTION ENZYMES ............................................................. 143 Recognition Site Features ............................................................. 143 Fragment Resolution .................................................................... 143 Sensitivity to Inhibitors and Stability ........................................... 143 Price and Availability ................................................................... 143 The Enzymes Selected for Forensic Analysis .............................. 143 REFERENCES .................................................................................. 145 8. PROBABILITY AND STATISTICAL ANALYSIS ...................... 149 STATISTICAL METHODOLOGY ................................................. 150 Random Sampling ........................................................................ 150 Data Presentation .......................................................................... 150
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