Table Of ContentM M B ™
ETHODS IN OLECULAR IOLOGY
Series Editor
John M. Walker
School of Life Sciences
University of Hertfordshire
Hatfield, Hertfordshire, AL10 9AB, UK
For further volumes:
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DNA Electrophoresis Protocols
for Forensic Genetics
Edited by
Antonio Alonso
Instituto Nacional de Toxicología y Ciencias Forenses, Servicio de Biología, Madrid, Spain
Editor
Antonio Alonso, Ph.D.
Instituto Nacional de Toxicología y Ciencias Forenses
Servicio de Biología, Madrid, Spain
[email protected]
ISSN 1064-3745 e-ISSN 1940-6029
ISBN 978-1-61779-460-5 e-ISBN 978-1-61779-461-2
DOI 10.1007/978-1-61779-461-2
Springer New York Dordrecht Heidelberg London
Library of Congress Control Number: 2011943292
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Preface
At present, most of the validated forensic DNA profi ling procedures are based on high-
resolution and high-throughput capillary electrophoresis separation and detection systems
of PCR amplicons obtained from DNA genomic markers with different inheritance pat-
terns (autosomal, Y-chromosome or X-chromosome linked, and mitochondrial DNA). A
high degree of standardization has been achieved in the fi eld of typing procedures for PCR
products obtained with fl uorogenic labeled primers, separated by capillary electrophoresis,
and detected by the fl uorescence emission induced by laser on a CCD camera. Current
systems have acquired a high degree of accuracy and precision in the identifi cation of allelic
variants (with a resolution of one base), and also an improvement in the sensitivity of PCR
amplicon detection (with reproducible signals even from picograms of DNA template).
Another advantage of current capillary electrophoresis systems (with 5–6 multichannel
detection technology) is a greater capacity for simultaneous detection of multiple markers.
It can be detected up to 15–26 short tandem repeats (STRs) multiallelic markers, perform
simultaneously about 50 biallelic single nucleotide polymorphisms (SNPs) or detect 4-dye-
terminator sequencing reactions in a single capillary electrophoresis injection.
Human forensic identifi cation is currently performed using commercial kits for STR
multiplexing. Basically there are two sets of standardized STR markers in the criminal DNA
databases around the world: the European standard set of 12 STR markers and the USA
CODIS standard of 13 STR markers. They form together a standard of 18 STR markers in
total. Development of mini-STR markers and their recent incorporation into commercial
kits have improved the application of these markers in severely degraded DNA samples.
Other source of human genetic variation used in forensic casework is the analysis of
sex chromosome markers. The Y-chromosome STR haplotypes are of special interest in
the specifi c analysis of the male component of DNA mixtures that are very frequently faced
by forensic labs in sexual assault cases. Another application of Y-STR haplotyping using a
commercial multiplex PCR system of 17 loci, is the identifi cation of human remains and
missing persons checked against the appropriate reference samples of paternal relatives. As
regards X-chromosome STR analysis, it provides complementary analysis in some kinship
defi ciency cases.
The automated sequence analysis of mitochondrial DNA hypervariable regions is espe-
cially important in the analysis of forensic samples negative for nuclear DNA, or when it is
necessary to identify human remains by comparing them with maternal relatives.
Multiplex PCR-CE assays of autosomal SNP markers have been developed for human
individualization of degraded samples and as a complementary tool in paternity testing.
The analysis of autosomal, Y-chromosome, and mtDNA SNPs with well-differentiated
allele or haplotype frequencies among global population-groups has been also applied to
the inference of likely ancestry.
DNA Electrophoresis Protocols for Forensic Genetics offers a comprehensive coverage of
the most modern current electrophoretic protocols and interpretation guidelines used in
forensic genetics for the analysis of amplifi ed human DNA fragments and DNA sequencing.
v
vi Preface
It includes protocols for profi ling of autosomal STRs (s ee Chapters 1 – 3 , and 1 3 – 15 ),
Y-STRs ( see Chapter 4 ), X-STRs ( see Chapter 5 ), autosomal SNPs and INDELS ( see
Chapters 6 – 8 , 10 ), Y-SNPs ( see Chapter 9 ), mtDNA-SNPs ( see Chapter 11 ), and mtDNA
hypervariable regions HV1 and HV2 (s ee Chapters 1 9 – 21 ).
Besides the use of DNA electrophoresis on different applications for human identifi ca-
tion (criminal investigations, kinship analysis, degraded samples, low template DNA, etc.)
the book also covers some interesting electrophoretic protocols for molecular identifi cation
of nonhuman species with interest in forensic botany (s ee Chapters 1 7 and 1 8 ), forensic
veterinary (s ee Chapters 1 6 , 22 , and 23 ), and microbial forensics (s ee Chapter 2 6 ).
Finally the book explores novel forensic applications of capillary electrophoresis to
target messenger RNA markers for body fl uid identifi cation (s ee Chapter 1 2 ), and some
forensic applications of microchip capillary electrophoresis (s ee Chapters 2 4 and 25 ).
DNA Electrophoresis Protocols for Forensic Genetics has been written by highly recog-
nized professionals and specialists from different forensic DNA laboratories around the
world dealing with casework and using validated technology and high quality standards.
This book was made possible, thanks to them.
Madrid, Spain Antonio Alonso
Contents
Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xi
PART I CURRENT PROTOCOLS ON CAPILLARY ELECTROPHORESIS
OF AMPLIFIED DNA FRAGMENTS
1 An Overview of DNA Typing Methods for Human Identification: Past,
Present, and Future . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
Robyn Thompson, Silvia Zoppis, and Bruce McCord
2 Capillary Electrophoresis and 5-Channel LIF Detection of a 26plex
Autosomal STR Assay for Human Identification. . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Carolyn R. Hill
3 Capillary Electrophoresis of MiniSTR Markers to Genotype Highly
Degraded DNA Samples. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31
Michael D. Coble
4 Interpretation Guidelines of a Standard Y-chromosome STR 17-plex PCR-CE
Assay for Crime Casework. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
Lutz Roewer and Maria Geppert
5 Capillary Electrophoresis of an X-Chromosome STR Decaplex
for Kinship Deficiency Cases. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57
Leonor Gusmão, Cíntia Alves, Iva Gomes, and Paula Sánchez-Diz
6 A 48-plex Autosomal SNP GenPlex™ Assay for Human Individualization
and Relationship Testing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Carmen Tomas, Claus Børsting, and Niels Morling
7 Typing of 49 Autosomal SNPs by Single Base Extension and Capillary
Electrophoresis for Forensic Genetic Testing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87
Claus Børsting, Carmen Tomas, and Niels Morling
8 A 34-plex Autosomal SNP Single Base Extension Assay for Ancestry
Investigations. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109
C. Phillips, M. Fondevila, and Maria Victoria Lareau
9 SNaPshot® Minisequencing Analysis of Multiple Ancestry-Informative
Y-SNPs Using Capillary Electrophoresis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127
Maria Geppert and Lutz Roewer
10 Capillary Electrophoresis of 38 Noncoding Biallelic Mini-Indels for Degraded
Samples and as Complementary Tool in Paternity Testing. . . . . . . . . . . . . . . . . . . . 141
Rui Pereira and Leonor Gusmão
vii
viii Contents
11 Capillary Electrophoresis of an 11-Plex mtDNA Coding Region SNP Single
Base Extension Assay for Discrimination of the Most Common Caucasian
HV1/HV2 Mitotype . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 159
Peter M. Vallone
12 Capillary Electrophoresis of a Multiplex Reverse Transcription-Polymerase
Chain Reaction to Target Messenger RNA Markers for Body Fluid
Identification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 169
Cordula Haas, Erin Hanson, and Jack Ballantyne
13 DNA Extraction from Aged Skeletal Samples for STR Typing by Capillary
Electrophoresis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 185
René Huel, Sylvain Amory, Ana Bilić, Stojko Vidović, Edin Jasaragić,
and Thomas J. Parsons
14 Interpretation Guidelines for Multilocus STR Forensic Profiles from Low
Template DNA Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
Zoran M. Budimlija and Theresa A. Caragine
15 Interpretation Guidelines for Mixed-STR Multilocus Electrophoretic Profiles. . . . . 213
Juan Antonio Luque
16 Capillary Electrophoresis Analysis of a 9-plex STR Assay for Canine
Genotyping . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 231
Barbara van Asch and António Amorim
17 Capillary Electrophoresis of DNA from Cannabis sativa for Correlation
of Samples to Geographic Origin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 241
Heather Miller Coyle
18 Capillary Electrophoresis of Multigene Barcoding Chloroplast Markers
for Species Identification of Botanical Trace Evidence. . . . . . . . . . . . . . . . . . . . . . . 253
Gianmarco Ferri, Beatrice Corradini, and Milena Alù
PART II CAPILLARY ELECTROPHORESIS AND DNA SEQUENCING
19 Capillary Electrophoresis of Big-Dye Terminator Sequencing Reactions
for Human mtDNA Control Region Haplotyping in the Identification
of Human Remains. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267
Marta Montesino and Lourdes Prieto
20 Capillary Electrophoresis of Human mtDNA Control Region Sequences
from Highly Degraded Samples Using Short mtDNA Amplicons . . . . . . . . . . . . . . 283
Odile M. Loreille and Jodi A. Irwin
21 Interpretation Guidelines of mtDNA Control Region Sequence
Electropherograms in Forensic Genetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 301
Manuel Crespillo Marquez
22 Capillary Electrophoresis of mtDNA Cytochrome b Gene Sequences
for Animal Species Identification. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 321
Adrian Linacre
23 Sequence Analysis of the Canine Mitochondrial DNA Control Region
from Shed Hair Samples in Criminal Investigations. . . . . . . . . . . . . . . . . . . . . . . . . 331
C. Berger, B. Berger, and W. Parson
Contents ix
PART III ADVANCES IN MICROCHIP CAPILLARY ELECTROPHORESIS
24 Integrated Sample Cleanup and Microchip Capillary Array Electrophoresis
for High-Performance Forensic STR Profiling. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 351
Peng Liu, Susan A. Greenspoon, Stephanie HI Yeung, James R. Scherer,
and Richard A. Mathies
25 Microchip Capillary Electrophoresis Protocol to Evaluate Quality
and Quantity of mtDNA Amplified Fragments for DNA Sequencing
in Forensic Genetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 367
Coro Fernández and Antonio Alonso
26 Microchip Capillary Electrophoresis of Multi-locus VNTR Analysis
for Genotyping of Bacillus Anthracis and Yersinia Pestis in Microbial
Forensic Cases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 381
Andrea Ciammaruconi
Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 391
