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ADVANCED TOPICS IN
FORENSIC DNA TYPING
 This work was funded in part by the National Institute of Justice (NIJ) through an interagency
agreement with the NIST Law Enforcement Standards Office. Points of view in this document
are those of the author and do not necessarily represent the official position or policies of the U.S.
Department of Justice or the National Institute of Standards and Technology. Certain commercial
equipment, instruments, and materials are identified in order to specify experimental procedures
as completely as possible. In no case does such identification imply a recommendation or endorse-
ment by the National Institute of Standards and Technology, nor does it imply that any of the
materials, instruments, or equipment identified are necessarily the best available for the purpose.
 ADVANCED
TOPICS IN
FORENSIC DNA
TYPING:
INTERPRETATION
JOHN M. BUTLER
National Institute of Standards and Technology
Gaithersburg, Maryland, USA

AMSTERDAM • BOSTON • HEIDELBERG • LONDON • NEW YORK

OXFORD • PARIS • SAN DIEGO • SAN FRANCISCO
SINGAPORE • SYDNEY • TOKYO
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(other than as may be noted herein).

Notices
Knowledge and best practice in this field are constantly changing. As new research and experience
broaden our understanding, changes in research methods, professional practices, or medical treatment
may become necessary.

Practitioners and researchers must always rely on their own experience and knowledge in evaluating
and using any information, methods, compounds, or experiments described herein. In using such
information or methods they should be mindful of their own safety and the safety of others, including
parties for whom they have a professional responsibility.

To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume
any liability for any injury and/or damage to persons or property as a matter of products liability,
negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas
contained in the material herein.

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Printed and bound in China

15 16 17 18 10 9 8 7 6 5 4 3 2 1
 Dedication

To my parents Doug and Marsha Butler

who instilled in me an important worldview of education and service
by which I interpret everything around me
and to Bruce Budowle
who helped me to become a better writer when I was a graduate student
and since then has challenged me to become a better scientist and a better man.
This page intentionally left blank
 Contents

Foreword ix
Introduction xi
Acknowledgments xvii
About the Author xix

I
DATA INTERPRETATION
1. Data Interpretation Overview 3
2. Data, Models, Thresholds 25
3. STR Alleles and Amplification Artifacts 47
4. STR Genotypes 87
5. STR Profiles 109
6. DNA Mixtures 129
7. Low-Level DNA and Complex Mixtures 159
8. Troubleshooting Data Collection 183

II
STATISTICAL INTERPRETATION
9. Statistical Interpretation Overview 213
10. STR Population Data Analysis 239
11. DNA Profile Frequency Estimates and Match Probabilities 281
12. DNA Mixture Statistics 309
13. Coping with Potential Missing Alleles 333
14. Relationship Testing: Kinship Statistics 349
15. Lineage Marker Statistics 403
16. Laboratory Reports: Communicating Results and Conclusions 445

vii
viii CONTENTS

Appendix 1: STR Allele Frequencies from U.S. Population Data 497

Appendix 2: NRC I & NRC II Recommendations 519
Appendix 3: DAB Recommendations on Statistics 529
Appendix 4: Worked Mixture Example 537

Index 569
 Foreword

Another book from the prolific writings of John Butler that keeps pace with the rapidly changing
world of DNA profiling in forensic science e it reminds me of the “Red Queen’s Race” in Lewis
Carroll’s Alice Through the Looking-Glass: “.it takes all the running you can do, to keep in the
same place. If you want to get somewhere else, you must run at least twice as fast as that!” Over
the past few years there have been major changes in DNA profiling technology and interpretation.
The introduction of more sensitive instrumentation, coupled with the introduction of modern multi-
plexed loci from the manufacturers that employ up to 24 loci with new biochemistry and detection
platforms, also realizes the dream of approaching the ultimate in sensitivity e it is almost a matter of
routine to detect DNA profiles from a handful of cells. Such advances are not without significant
challenges, particularly in the area of interpretation of the evidence. Fortunately there have been
significant advances in this area too, although the complete adoption by the community is yet to
be realized.
The book is divided into 16 chapters and 4 appendices that describe the “state of the art” and
beyond. Starting with an overview of data interpretation, subsequent chapters lead onto a discussion
of the characterization of DNA profiles in terms of heterozygote balance, stutter, artifacts, and muta-
tion and stochastic effects. There is an introduction to the phenomenon of allele drop-out, a character-
istic of low-template DNA that results in “false homozygotes.” To interpret DNA profiles, a system of
thresholds (e.g. stochastic threshold, stutter threshold, limit of detection) has evolved. However,
there are limitations and risks to consider. There is an extensive discussion on the deliberation of
the scientific societies that is linked to an outline of the steps to interpret mixtures, before moving
onto challenging “complex DNA profiles,” i.e. those profiles that are mixtures of two or more contrib-
utors, where allele drop-out, secondary transfer, and contamination are additional complications.
There has been a fundamental shift by the forensic community towards the analysis of these kinds
of profiles. In one laboratory we are informed that the proportion of submissions of samples with
<100 pg DNA has increased from 19% in 2004 to 45% in 2008. Chapters 9e13 are devoted to the inter-
pretation of evidence complemented by Appendix 1, which has a complete list of STR loci used in the
U.S. and their respective allele frequencies. There is a comprehensive review of statistical theory and
methods e this leads to a complete description of various programmed solutions that have recently
evolved to interpret “complex DNA profiles.” Appendix 4 has a worked example, concisely written
by Mike Coble, that further explains the rationale, theory, and benefits of the practical use of prob-
abilistic methods. The reader is able to use the book as a handy one-stop guide to everything that
is currently available (and where to find out more information).
Chapter 14 turns to relationship testing, providing an outline of the theory, a summary of recom-
mendations from the scientific societies, a list of available software, along with an outline of the use of
high-density SNP arrays to identify distant relationships. The penultimate Chapter 15 deals with
lineage markers, including mitochondrial DNA, and Y-chromosomal and X-chromosomal DNA.

ix Copyright Ó 2015 Elsevier Inc. All rights reserved.

x FOREWORD

These markers are useful in missing persons, disaster victim identification, and other complex
inquires.
Finally, Chapter 16 explores the most contentious area in forensic science, namely, the interpreta-
tion of the evidence. It isn’t just the fact of the DNA profile to consider. With the introduction of
methods that are ever more sensitive, the focus turns to how and when the DNA profile was trans-
ferred. Is it possible that a contamination event, or secondary transfer is possible? How can cognitive
(psychological) bias be avoided in reporting? A useful way to think about evidence is described by
the “hierarchy of propositions” e can we ascribe a “source,” such as blood, or an “activity,” such as
stabbing, to the DNA profiling evidence? To prepare this chapter, John Butler has consulted widely
with practitioners, gathering their views, cross-referenced to historical and recent deliberations of the
scientific societies and advisory boards (Appendices 2 and 3), distilling the information into a treatise
on how to write a report for the court.
This book complements John Butler’s previous works with the most comprehensive and
up-to-date text of its kind. As such, it will readily be adopted by the forensic community as the defin-
itive guide to the galaxy of forensic DNA-typing technologies.

Peter Gill, Ph.D.

April 2014
 Introduction

“Writing is thinking. To write well is to think clearly. That’s why it’s so hard.”
David McCullough (NEA 2003)

The third edition of Forensic DNA Typing has been divided up into three volumes: a basic
volume for students and beginners in the field and two advanced volumes for professionals/
practitioners who may be interested in more detail. The basic volume was released in September
2009 (with a publication date of 2010) and is entitled Fundamentals of Forensic DNA Typing. The first
advanced volume, Advanced Topics in Forensic DNA Typing: Methodology, was released in August
2011 (with a 2012 publication date). The present book, Advanced Topics in Forensic DNA Typing:
Interpretation, is intended as volume 3 of the third edition, with a focus on data interpretation
and statistical analysis.
Several reasons exist for dividing the material. First and foremost, people use books more
frequently if they are less bulky. I have heard from more than one colleague at conferences that
they prefer to carry the smaller first edition with them to court or other teaching situations. Second,
by having multiple books, each volume can be focused on its intended audience rather than trying to
be all things to all readers. Third, the books will enable both undergraduate and graduate studies,
with each building upon the previous volumes.
There is only minor overlap in subject matter among the various volumes. The basic Fundamentals
volume contains the simpler “starter” information, while most of the “updates” to the field are found
in the Advanced Topics volumes. It is my intention that the three volumes together provide a compre-
hensive view of the current state of forensic DNA analysis. With a field advancing as rapidly as
forensic DNA typing is, this is a challenge.
The present book has been divided into two primary sections: data interpretation (Chapters 1e8)
and statistical interpretation and reporting (Chapters 9e16). The first part covers data analysis
and factors impacting a DNA profile, while the second part examines evidence evaluation and
interpretation e essentially what information exists in a DNA profile and what does this information
mean in the context of variability expected from the DNA results obtained. In this edition, I again
utilize Data, Notes, and Applications (D.N.A.) Boxes to cover specific topics of general interest, to
review example calculations, or to cover a topic that serves to highlight information needed by
a DNA analyst.

NEW MATERIAL IN THIS VOLUME

Advanced Topics in Forensic DNA Typing: Interpretation is substantially enhanced with additional
information beyond what was available in the second edition of Forensic DNA Typing, which was

xi
xii INTRODUCTION

completed in June 2004. Much has happened to advance our understanding of DNA evidence inter-
pretation in the past decade.
I am grateful to have had three years between writing the Methodology and this Interpretation
volume. During this additional time, I have learned a great deal as I have responded to numerous
questions from forensic scientists around the world via email or in person. I have also had the priv-
ilege of preparing and presenting hundreds of slides on DNA mixture interpretation. Materials from
these training workshops are available at the NIST website (NIST 2014a). In addition, recent publi-
cations have provided new research and perspectives on DNA interpretation. For example, the Inter-
national Society for Forensic Genetics DNA Commission published recommendations on DNA
mixtures and probabilistic approaches in the December 2012 issue of Forensic Science International:
Genetics.
As with previous books, a fairly comprehensive list of references is included at the end of each
chapter that serves as a foundation for citations in the chapter as well as a launching point where
interested readers can go for additional information. More than 2,000 references are provided,
enabling readers to expand their study beyond the information contained between the covers of
this book. References to journal articles include titles to enhance value.
More than 80% of this book is completely new. Figures, tables, and D.N.A. Boxes have been
created (>200 in total) to help illustrate the principles being taught. Since I do not now write or
review laboratory reports, I sought and received valuable input from more than a dozen laboratory
analysts, laboratory directors, police investigators, lawyers (prosecution and defense), and private
consultants who regularly review laboratory reports. Their collective wisdom and insights are
captured in Chapter 16. Numerous others have contributed to information contained in this volume,
and their input is gratefully appreciated and acknowledged.
Throughout this book I have prepared teaching examples with the STR locus D18S51, which is one
of the original U.S. core markers and also is used in Europe, China, Australia, and elsewhere around
the world. I have endeavored to mind my Ps and Qs in using a consistent allele nomenclature in
statistical equations. In some areas, material in this book may not be as advanced as readers might
like. There is, however, enough information to help provide a bridge to more detailed work already
available or soon to be available from authors like John Buckleton and Peter Gill.
At points throughout the text, I quote from the 2010 SWGDAM Interpretation Guidelines for
Autosomal STR Typing by Forensic DNA Testing Laboratories. Although I chaired the group that
prepared these guidelines, the opinions expressed in this book are mine alone and in no way should
be thought of as the official opinion of SWGDAM or the SWGDAM Autosomal STR Interpretation
Committee.

OVERVIEW OF BOOK CHAPTERS

Chapters 1 to 8 cover data interpretation. Chapter 1 provides an overview and perspective on
principles, protocols, and practice in an effort to understand the “why,” “what,” and “how” of
forensic DNA typing. Autosomal and Y-chromosome STR loci and kits are reviewed. Chapter 2
describes data generation by the Applied Biosystems Genetic Analyzers as well as introducing
the role of statistical models in setting thresholds such as the analytical threshold. Chapter 3
covers issues surrounding measurement of STR alleles and distinguishing alleles from artifacts
such as stutter products. Chapter 4 reviews peak height ratios (heterozygote balance) and null
 INTRODUCTION xiii
alleles that can impact STR genotypes. Stochastic thresholds and allele drop-out are introduced.
Chapter 5 focuses on evaluation of multi-locus STR profiles, tri-allelic patterns, and issues with
amelogenin as a sex-typing marker. Chapter 6 considers DNA mixtures and steps for interpreting
them, including determining the number of contributors. Chapter 7 explores complex DNA
mixture issues, including allele sharing and allele drop-out due to low-level DNA contributors.
Chapter 8 seeks to inform readers regarding principles and processes involved with capillary elec-
trophoresis so that high-quality data may be obtained and common problems avoided. It
discusses how troubleshooting improves as close attention is paid to details in the laboratory
data produced.
Chapters 9 to 16 involve statistical interpretation and reporting. Chapter 9 introduces the role of
statistics in forensic DNA analysis with a review of the individuals who have influenced the field.
The laws of probability are introduced along with likelihood ratios and Bayesian versus frequentist
approaches to statistics. Chapter 10 describes principles of population genetics and how population
data are used to estimate STR profile frequencies. Chapter 11 covers approaches and assumptions
made with generating STR profile frequency estimates. Chapter 12 involves DNA mixture statistics
and contrasts combined probability of inclusion methods with likelihood ratio calculations.
Chapter 13 deals with situations where allele drop-out from stochastic effects with low-level DNA
present challenges in comparing evidentiary profiles to reference profiles. Chapter 14 reviews kinship
analysis used in relationship testing. Paternity testing, mutations, and disaster victim identification
are among the topics discussed. Chapter 15 addresses Y-chromosome, X-chromosome, and mito-
chondrial DNA lineage markers and how their different genetic transmission influences statistical
interpretation of results. Finally, Chapter 16 covers laboratory reports and the importance of effec-
tively communicating results and conclusions. Input from more than a dozen laboratory report
providers and users enhanced this material.

Appendices
There are four appendices at the back of the book that provide helpful supplemental material:
• Appendix 1 provides U.S. population data in the form of STR allele frequencies based on studies
performed at NIST. This information is utilized in worked examples throughout the book.
• Appendix 2 lists the recommendations made by both National Research Council reports (NRC I
and NRC II) in their 1992 and 1996 publications entitled “DNA Technology in Forensic Science”
and “The Evaluation of Forensic DNA Evidence.”
• Appendix 3 contains the FBI’s DNA Advisory Board recommendations on statistics that were
released in February 2000 to provide a historical perspective.
• Appendix 4 is a DNA mixture example prepared by Dr. Mike Coble, a valued colleague within the
Applied Genetics Group at the National Institute of Standards and Technology.
A brief “cross-walk” of major topics covered across the various editions of Forensic DNA Typing is
shown in Table I.1 with chapters (Ch.) and appendices (App.) indicated. In a few cases, information
was limited to a single D.N.A. Box.
In my Interpretation book, I have included information from the latest articles as well as insights I
have gained over the past two decades of working in the field. Writing these books on forensic
DNA typing has been richly rewarding as I must carefully think through each issue and decide
how to best address it. As David McCullough points out in the quote at the beginning of this
xiv INTRODUCTION

TABLE I.1 “Cross-Walk” of Major Topics

3rd edition, 3rd edition, 3rd edition,

volume 1 volume 2 volume 3

1st edition 2nd edition Fundamentals Advanced Topics: Advanced Topics:

Topic (2001) (2005) (2010) Methodology (2012) Interpretation (2015)

Amelogenin Ch. 5 Ch. 5 Ch. 8 Ch. 5 Ch. 5

Capillary Ch. 9 & 11 Ch. 12 & 14 Ch. 9 Ch. 6 Ch. 8
electrophoresis
Data interpretation Ch. 6 & 13 Ch. 6 & 15 Ch. 10 e Ch.1e8

Disaster victim Ch. 17 Ch. 24 Ch. 17 Ch. 9 Ch. 14

identification
DNA basics Ch. 2 Ch. 2 Ch. 2 e e
DNA databases Ch. 16 Ch. 18 Ch. 12 Ch. 8 e
DNA extraction Ch. 3 Ch. 3 Ch. 5 Ch. 2 e

DNA quantitation Ch. 3 Ch. 3 Ch. 6 Ch. 3 e

Expert witness e e e Ch. 18, App. 4 e
testimony
Familial searching e e Ch. 12 (p. 282) App. 2 D.N.A. Box 14.9

FBI Quality App. 3 App. 4 e e e

Assurance Standards (1998/99) (1998/99)

FMBIO gel imaging Ch. 12 Ch. 14 D.N.A. Box 9.2 e e

system

Glossary e e App. 1 e e
History of DNA Ch. 1 Ch. 1 Ch. 1 & 3 e e

Kinship analysis e Ch. 23 Ch. 17 e Ch. 14

Low copy number e Ch. 7 Ch. 14 Ch. 11 Ch. 7
DNA testing
Match probability e Ch. 21 Ch. 11 e Ch. 11
calculations

Mixtures Ch. 7 Ch. 7 Ch. 14 e Ch. 6 & 7, App. 4

Mixture statistics e Ch. 22 D.N.A. Box 14.2 e Ch. 12 & 13
Mitochondrial DNA Ch. 8 Ch. 10 Ch. 16 Ch. 14 Ch. 15

New technologies Ch. 15 Ch. 17 Ch. 18 Ch. 17 e

Non-human DNA Ch. 8 Ch. 11 Ch. 15 Ch. 16 e
Null alleles Ch. 6 Ch. 6 D.N.A. Box 10.3 Ch. 5 Ch. 4

PCR Ch. 4 Ch. 4 Ch. 7 Ch. 4 e

 INTRODUCTION xv
TABLE I.1 “Cross-Walk” of Major Topics (cont'd)
3rd edition, 3rd edition, 3rd edition,
volume 1 volume 2 volume 3

1st edition 2nd edition Fundamentals Advanced Topics: Advanced Topics:

Topic (2001) (2005) (2010) Methodology (2012) Interpretation (2015)

Population data e Ch. 20 Ch. 11 e Ch. 10

Report writing e e e e Ch. 16

Sample collection Ch. 3 Ch. 3 Ch. 4 Ch. 1 e

SNP testing Ch. 8 Ch. 8 Ch. 15 Ch. 12 e

Statistics & e Ch. 19 App. 3 e Ch. 9
probability
STR alleles App. 1 App. 1 e App. 1 Ch. 3

STR kits Ch. 5 Ch. 5 Ch. 8 Ch. 5 Ch. 1

STR markers Ch. 5 Ch. 5 Ch. 8 Ch. 5 Ch. 1
Stutter products Ch. 6 Ch. 6 Ch. 10 e Ch. 3
Thresholds e e Ch. 10 e Ch. 2

Tri-allelic patterns Ch. 6 Ch. 6 D.N.A. Box 10.2 e Ch. 5

Validation Ch. 14 Ch. 16 Ch. 13 Ch. 7 Ch. 5

Variant alleles Ch. 6 Ch. 6 D.N.A. Box 10.1 App. 1 Ch. 3
X-STRs e e e Ch. 15 Ch. 15

Y-STRs Ch. 8 Ch. 9 Ch. 16 Ch. 13 Ch. 15

Introduction, writing is hard work. This hard work has been beneficial to my personal learning but
comes at the price of significant time away from my family and other responsibilities. I am grateful
for the support of others, especially my wife, who have permitted me the time needed to complete
this book.
My father, Doug Butler, has written about a dozen textbooks and spent his career teaching and
helping to shape his profession. He is an amazing teacher because of his dedication to learning
and excelling at the highest level. Recently when I told him some of the things I was learning while
working on this book and other presentations that I was giving to the forensic DNA community, he
shared an important lesson: “You never really learn anything until you have to teach it to someone
else.” While I hope the information in this book helps the field, I know that I have been the main bene-
ficiary of this effort in terms of what I have learned during the process of trying to teach the concepts
to others.
The pressure to carefully craft each phrase so that my words are less likely to be misunderstood
has increased with the widespread use of my books, particularly in courts of law. I do not take lightly
the opportunity to share my thoughts and perspective in this book. While I have benefited from
discussions and input from many people, I alone am responsible for the content. My goal in
xvi INTRODUCTION

preparing training materials on the topic of forensic DNA has always been to be on the side of good
science that is well-documented and appropriately applied to benefit the forensic science community.
I hope that this book contributes to that goal.

References
National Endowment for the Arts (NEA). Awards & Honors, 2003 Jefferson Lecturer. http://www.neh.gov/about/awards/
jefferson-lecture/david-mccullough-interview. Accessed Apirl 2, 2014.
NIST (2014a). DNA Mixture Interpretation. http://www.cstl.nist.gov/strbase/mixture.htm. Accessed Apirl 2, 2014.
 Acknowledgments

I express a special thanks to colleagues and fellow researchers who kindly provided important infor-
mation and supplied some of the figures for this book or previous editions of Forensic DNA Typing. The
list continues to grow. These individuals include Ricky Ansell, Michael Baird, Susan Ballou, Brad
Bannon, Leslie Biesecker, Martin Bill, Erica Butts, Lisa Calandro, Theresa Caragine, George Carmody,
Ranajit Chakraborty, Tim Clayton, Mike Coble, Robin Cotton, Cecelia Crouse, Amy Decker,
Christopher Duby, David Duewer, Dan Ehrlich, Nicky Fildes, Lisa Forman, Ron Fourney, Lee Fraser,
Dave Gillespie, Catherine Grgicak, Richard Guerrieri, Ravi Gupta, Chip Harding, Doug Hares, Bruce
Heidebrecht, Mary Herdman, Becky Hill, Debbie Hobson, Bill Hudlow, Ted Hunt, Alice Isenberg,
Dennis Kilcoyne, Margaret Kline, Sonja Klein, Ken Konzak, Carll Ladd, Steve Lee, Dina Mattes, Bruce
McCord, Terry Melton, Ruth Montgomery, Niels Morling, Steven Myers, Steve Niezgoda, Kristen
Lewis O’Connor, Antonio Possolo, Mecki Prinz, George Riley, Norah Rudin, Jeff Sailus, Thomas
Schnibbe, Richard Schoske, Jim Schumm, Scott Scoville (and the Orange County DA’s DNA Unit),
Bob Shaler, Michelle Shepherd, Gary Sims, Melissa Smrz, Amanda Sozer, Jill Spriggs, Mark Stolorow,
Kevin Sullivan, Lois Tully, Pete Vallone, Ray Wickenheiser, and Charlotte Word.
I am indebted to the dedicated human identity project team members, past and present, who have
worked with me at the National Institute of Standards and Technology: Jill Appleby, Erica Butts,
Mike Coble, Amy Decker, David Duewer, Becky Hill, Kevin Kiesler, Margaret Kline, Kristen Lewis
O’Connor, Jan Redman, Dennis Reeder, Patti Rohmiller, Christian Ruitberg, Richard Schoske, and
Pete Vallone. It has been a pleasure to work with such supportive and hard-working scientists. Since
I moved into a new role at NIST in April 2013, I miss the daily interaction with the Applied Genetics
Group.
Several other people deserve specific recognition for their support of this endeavor. The informa-
tion reported in this book was in large measure made possible by a comprehensive collection of refer-
ences on the STR markers used in forensic DNA typing. For this collection, now numbering more
than 3,500 references, I am indebted to the initial work of Christian Ruitberg for tirelessly collecting
and cataloging these papers and the steady efforts first of Jan Redman and then Patti Rohmiller to
regularly update this STR reference database. A complete listing of these references may be found
at the NIST website (NIST 2014b).
My wife Terilynne, who carefully reviewed the manuscript and made helpful suggestions, was
always a constant support in the many hours that this project took away from our family. As the
initial editor of all my written materials, Terilynne helped make the book more coherent and read-
able. In addition, David Duewer and Katherine Sharpless provided a fine technical review of my
two previous books as well as this one. The support of NIST management, especially Laurie Locascio,
Mike Tarlov, Richard Cavanagh, and Willie May, made completion of this book possible.

Reference
NIST (2014b). STRs and DNA Typing. http://www.cstl.nist.gov/strbase/str_ref.htm. Accessed April 2, 2014.

xvii
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 About the Author

John Marshall Butler grew up in the U.S. Midwest and, enjoying science and law, decided to
pursue a career in forensic science at an early age. After completing an undergraduate education
at Brigham Young University in chemistry, he moved east to pursue graduate studies at the Univer-
sity of Virginia. While a graduate student, he enjoyed the unique opportunity of serving as an FBI
Honors Intern and guest researcher for more than two years in the FBI Laboratory’s Forensic Science
Research Unit. His Ph.D. dissertation research, which was conducted at the FBI Academy in
Quantico, Virginia, involved pioneering work in applying capillary electrophoresis to STR typing.
After completing his Ph.D. in 1995, Dr. Butler obtained a prestigious National Research Council post-
doctoral fellowship to the National Institute of Standards and Technology (NIST). While a postdoc at
NIST, he designed and built STRBase, the widely used Short Tandem Repeat Internet Database
(STRBase 2014) that contains a wealth of standardized information on STRs used in human identity
applications. He worked for several years as a staff scientist and project leader at a California startup
company named GeneTrace System developing rapid DNA analysis technologies involving time-
of-flight mass spectrometry. In the fall of 1999, he returned to NIST to lead their efforts in human
identity testing with funding from the National Institute of Justice. He served as leader of the Applied
Genetics Group from 2008 to 2013.
Dr. Butler is a NIST Fellow and Special Assistant to the Director for Forensic Science. He is a regular
invited guest of the FBI’s Scientific Working Group on DNA Analysis Methods (SWGDAM) and
served for many years on the Department of Defense Quality Assurance Oversight Committee for
DNA Analysis. Following the terrorist attacks of 11 September 2001, he aided the DNA identification
efforts and served as part of the distinguished World Trade Center Kinship and Data Analysis Panel
(WTC KADAP). He is a member of the International Society of Forensic Genetics and the American
Academy of Forensic Sciences. Dr. Butler serves as an Associate Editor for Forensic Science
International: Genetics and is on the editorial board for the Journal of Forensic Sciences.
Dr. Butler has received numerous awards, including the Presidential Early Career Award for
Scientists and Engineers (2002), the Department of Commerce Silver Medal (2002) and Gold Medal
(2008), the Arthur S. Flemming Award (2007), the Edward Uhler Condon Award (2010), Brigham
Young University’s College of Physical and Mathematical Sciences Honored Alumnus (2005), and
the Scientific Prize of the International Society of Forensic Genetics (2003).
He has more than 150 publications describing aspects of forensic DNA testing and is one of the
most prolific active authors in the field, with articles appearing regularly in every major forensic
science journal. In 2011, ScienceWatch, which monitors the number of citations by other scientists
that authors receive for their research work, reported that Dr. Butler was the #1 world-wide high-
impact author in legal medicine and forensic science over the decade of 2001e2011.
Dr. Butler has been an invited speaker to numerous national and international forensic DNA meet-
ings and in the past few years has spoken in Argentina, Australia, Austria, Belgium, Brazil, Canada,

xix
xx ABOUT THE AUTHOR

China, Cyprus, Denmark, England, France, Germany, Israel, Japan, Korea, Mexico, The Netherlands,
Poland, Portugal, Sweden, and Taiwan. Much of the content in this book has come from the NIST
Applied Group’s research efforts over the past two decades. In addition to his busy scientific career,
he and his wife serve in their community and church and are the proud parents of six children, all of
whom have been proven to be theirs through the power of DNA typing.

Reference
STRBase (2014). http://www.cstl.nist.gov/strbase. Accessed April 2, 2014.
 S E C T I O N I

DATA INTERPRETATION
This page intentionally left blank
 C H A P T E R

1
Data Interpretation Overview
“We see the world, not as it is, but as we are e or, as we are conditioned to see it.”
Stephen R. Covey (The 7 Habits of Highly Effective People, p. 28)

PURPOSE OF THIS BOOK

This book is primarily intended for DNA analysts or those trying to understand what a DNA
analyst does in his or her review of forensic DNA data that was obtained by polymerase chain reaction
(PCR) amplification and short tandem repeat (STR) typing via capillary electrophoresis (CE). A DNA
analyst, according to the FBI Quality Assurance Standards (QAS) that govern U.S. laboratories, is an
individual who “conducts and/or directs the analysis of forensic samples, interprets data and reaches
conclusions” (QAS 2011, definitions). Many laboratories employ technicians to perform the analytical
techniques required to obtain a DNA profile from a biological sample e typically under the supervi-
sion of a trained and qualified analyst. However, as noted by the QAS, “technicians do not interpret
data, reach conclusions on typing results, or prepare final reports” (QAS 2011, definitions). Thus, there
is an expectation that DNA analyst training involves developing an understanding and mastery of
data interpretation as well as report writing and statistical analysis used in reaching conclusions on
typing results.
The general steps and workflow involved in forensic DNA typing are illustrated in Figure 1.1.
The companion volume to this book entitled Advanced Topics in Forensic DNA Typing: Methodology

Understanding
Results Obtained
Gathering the Data & Sharing Them
Collection/Storage/ Extraction/ Amplification/ Separation/
Data Stats Report
Characterization Quantitation Marker Sets Detection

Advanced Topics: Methodology Interpretation

Advanced Topics: Interpretation

FIGURE 1.1 Steps involved in the overall process of forensic DNA typing. This book focuses on understanding the data
through data interpretation and statistical interpretation.

Advanced Topics in Forensic DNA Typing: Interpretation

http://dx.doi.org/10.1016/B978-0-12-405213-0.00001-4 3 2015 Published by Elsevier Inc.
4 1. DATA INTERPRETATION OVERVIEW

(Butler 2012) covered many aspects of gathering the data used in DNA testing. Picking up where that
book left off, the purpose of this book, Advanced Topics in Forensic DNA Typing: Interpretation, is to
help readers understand data obtained from the STR typing process, with a focus on interpreting
and reporting results. Data interpretation is covered in Chapters 1 through 8 where we address
the question, “What are the data obtained from a set of samples?” Statistical interpretation is
reviewed in Chapters 9 through 15 to help discuss, “How significant are the data?” Chapter 16 fo-
cuses on drawing conclusions and report writing to assess, “What do the data mean when compar-
isons are made between evidentiary and reference sample results?”
Everyone may think that their way of DNA analysis is correct. However, misinterpretations of
some fundamental principles have given rise to a variety of approaches being undertaken in labs
today, some of which are not optimal, or even border on being incorrect for certain scenarios of
use. Unfortunately, often times the approaches taken for interpretation are subjective, and therefore
become the weakest part of the overall DNA typing process. I have written this book because I
believe that a better understanding of fundamental principles will aid consistency and quality of
work being performed in forensic DNA laboratories around the world.
In February 2009, the U.S. National Academy of Sciences released a report entitled “Strengthening
Forensic Science in the United States” (NAS 2009). The report emphasized that good (forensic) science
includes: (1) valid and reliable methodologies, and (2) practices that minimize the threat of bias in
data interpretation. My Methodology volume demonstrates that valid and reliable methodologies
can be achieved with forensic DNA typing. This Interpretation volume seeks to help minimize the
threat of bias in data interpretation.
Good science takes time and effort to do well. It is worth noting that some measurements and in-
terpretations are more reliable than others. Hence, uncertainty in measurements and interpretation
should be reflected in the reports generated in a forensic case investigation. As will be described
throughout this book, it is important that assumptions made during the interpretation process be
documented and conveyed as clearly as possible. This documentation will aid those individuals
reviewing the lab report to appropriately assess the results obtained and the conclusions drawn. It
is important for analysts to offer what they know from the data obtained in a case in a fashion
that is as clear and unbiased as possible.
That being said, I recognize that there are two areas of forensic DNA interpretations that are partic-
ularly challenging: (1) low-level DNA samples where sensitivity is an issue, and (2) complex mixtures
where specificity is an issue. In other words, how much DNA is needed to obtain a reliable result and
how well can the number of contributors to a sample be estimated to limit the uncertainty or ambi-
guity in the conclusions drawn. Chapters 7 and 13 will discuss some potential approaches to
handling difficult interpretations. Unfortunately, in many situations involving complex results where
uncertainty in the interpretation is large, the only scientifically responsible conclusion is “inconclu-
sive” to avoid the chance of inappropriately including or excluding a potential contributor from
an evidentiary result.

THE INTERPRETATION PROCESS

If the companion volume entitled Advanced Topics in Forensic DNA Typing: Methodology begins with
an evidentiary biological sample from a crime scene, then this book begins with a computer file. This
computer file contains data points corresponding to time and fluorescence intensity at various

I. DATA INTERPRETATION
 THE INTERPRETATION PROCESS 5
wavelengths of light that represent the digital signature of a DNA profile. When these data points are
plotted with time on the x-axis and fluorescence intensity on the y-axis, an electropherogram is
created. This electropherogram, sometimes referred to as an EPG or e-gram, is then evaluated using
STR genotyping software to produce a final results table representing the biological sample’s DNA
profile.
An overview of the components and processes involved in data interpretation are illustrated in
Figure 1.2. A sample data file contains time and fluorescence information for a PCR-amplified sample
along with an internal size standard. The sample data file, which has a file extension of .fsa or .hid, is
loaded into genotyping software along with an allelic ladder data file containing the same internal
size standard to enable the sample and allelic ladder results to be correlated. Along with the allelic
ladder sample, STR kit manufacturers provide a computer file specific for each STR kit containing
bins (that define the allele repeat number for each STR locus) and panels (that define the STR loci pre-
sent in the kit). When combined with the allelic ladder data file, bins and panels provide genotyping
software with the capability to transform DNA size information into an STR allele repeat number for
each observed peak.

Laboratory Protocols to Aid Interpretation

Laboratory protocols, which are often referred to as standard operating procedures (SOPs), are
step-by-step instructions used to provide a consistent framework to gather and interpret information
from analyzed samples. As part of a quality assurance system, accredited forensic laboratories will
have written SOPs. Laboratory personnel are trained to understand and follow their laboratory-
specific SOPs.
As will be described in more detail in Chapter 2, laboratories define parameters as part of their
SOPs that act as thresholds within the genotyping software to filter information and aid analyst de-
cisions that are made in determining the final sample DNA profile. These SOPs should be created
based on validation data and then verified to work properly with control samples before being
put into routine use.
A primary purpose of SOPs is to provide consistent results across DNA analysts within a labora-
tory as well as across cases analyzed by the same DNA analyst. The hope is that by following well-
defined directions in a laboratory’s SOP, the same result can be obtained on a particular DNA sample
by any qualified analyst or data reviewer.

Overview of Data Interpretation Process FIGURE 1.2 Overview of DNA

Allelic Ladder Data File
(with internal size standard)
Bins & Panels
Sample Data File Genotyping
(with internal size standard) Software
Laboratory SOPs
with parameters/thresholds
established from validation studies
interpretation process illustrating
that sample data files, at least one
allelic ladder data file, and informa-
tion from laboratory SOPs are
entered into genotyping software.
Analyst or Sample Analysts (or expert system software)
Expert System DNA Profile review the information from the
Decisions software to produce the final sample
DNA profile.

I. DATA INTERPRETATION
6 1. DATA INTERPRETATION OVERVIEW

Decisions during Data Interpretation

An analyst must make decisions about whether electrophoretic data from an evidentiary or a refer-
ence sample represent peaks or noise, whether peaks are alleles or artifacts, whether alleles can be
confidently paired to form genotypes, whether genotypes from individual loci can be combined to
create a contributor profile, whether the data are too weak or too complex to be reliably interpreted,
and if overall data quality is appropriate for obtaining reliable results. Table 1.1 correlates the discus-
sion of further details on these decisions with the various chapters in the first half of this book.
A DNA profile produced from the evidentiary sample, often referred to as the question (Q) sam-
ple, is then compared to a reference, or known (K) sample, which must also undergo data analysis
and the same interpretation decision process. Reference samples may come directly from a suspect
or indirectly from a DNA database search of previous offenders. Fortunately, expert system software
programs have been validated and implemented in many labs to help rapidly evaluate single-source
samples (see Butler 2012, Table 8.4).
Following comparison of the Q and K sample profile results, conclusions are drawn regarding a
potential match or not, and a report is written (see Chapter 16). If there is deemed to be a match
(or some kind of kinship association) between the Q and K samples, then statistical interpretation
is performed to estimate the weight-of-evidence. Chapters 9 through 15 describe approaches for
statistical interpretation and issues involved. A summary of the steps and decisions in STR data
interpretation are illustrated in Figure 1.3.

The DNA Profile Computer File

The computer file extension for a DNA result produced by an Applied Biosystems Genetic
Analyzer will be either .fsa or .hid depending on the instrument used to collect data from separated
components of PCR-amplified STR markers. The .fsa (fragment size analysis) files are produced with
ABI 310, 3100, 3130, 3700, and 3730 series CE instruments, while the .hid (human identity) files are
produced with ABI 3500 series CE systems.

TABLE 1.1 Information Flow in the Data Interpretation Process Correlated with Chapters in This Book

Chapter Input Information Decision to be made How decision is made

2 Data file Peak or Noise Analytical threshold

3 Peak Allele or Artifact Stutter threshold; precision sizing bin

4 Allele Heterozygote or Homozygote or Allele(s) Peak heights and peak height ratios;
missing stochastic threshold

5 Genotype/full profile Single-source or Mixture Numbers of peaks per locus

6 Mixture Deconvolution or not Major/minor mixture ratio
7 Low level DNA Interpret or not Complexity/uncertainty threshold

8 Poor quality data Replace CE components (buffer, polymer, Review size standard data quality with
array) or call service engineer understanding of CE principles

I. DATA INTERPRETATION
 THE INTERPRETATION PROCESS 7

Steps in DNA Interpretation FIGURE 1.3 Steps in DNA interpreta-

Match probability
tion. The evidentiary (Q) sample and
Question sample reference (K) sample are processed from
peaks to profile and then compared. If Q
Weight
Peak Allele Genotype Profile and K match, then a match probability is
of
(vs. noise) (vs. artifact) (allele pairing) (genotype combining) computed to assess the weight of this
Evidence
evidence. Finally, a report is written
Known sample describing the results obtained. A separate
technical review by another analyst in the
originating laboratory is performed prior
to the casework report being finalized and
Report Written released by the laboratory.
& Reviewed

As described in D.N.A. Box 1.1, the .fsa files are in a binary file format known as ABIF (Applied
Biosystems, Inc. Format) and are similar to Tag Image File Format (TIFF), which has been used for
graphics files (Applied Biosystems Genetic Analysis Data File Format 2009). Although we will
focus on the electronic information used to create a DNA profile, it is worth noting that other
diagnostic information, such as laser power and run current, is also stored in the computer file
during data collection; this information can be helpful in troubleshooting efforts described in
Chapter 8.
During the process of data analysis and genotyping, information from the .fsa or .hid data
file is converted from time (scan) points to DNA size relative to an internal size standard
and then to an STR allele call relative to STR typing kit-specific bins and panels and allelic
ladders.

Software for Analysis of DNA Profile Computer Files

Sophisticated software has been developed to take sample electrophoretic data rapidly through
the STR genotyping process (Ziegle et al. 1992). Life Technologies/Applied Biosystems (Foster
City, CA), which manufactures the Genetic Analyzer CE instruments used in forensic DNA lab-
oratories, supplies software for processing the .fsa or .hid files generated by their CE instruments.
This software enables peaks to be defined and STR alleles designated using kit-specific allelic
ladders and bins and panels. GeneScan and Genotyper software programs were used originally
with early Mac and NT versions of data collection from the ABI 310 and 3100 series instruments.
In more recent years, GeneMapperID v3.2 and GeneMapperID-X v1.1 or 1.2 have replaced
GeneScan/Genotyper functions (in 2012, GeneMapperID-X v1.4 expanded data analysis capabil-
ities to 6-dyes).
GeneMarkerHID software (Holland & Parson 2011) from Soft Genetics (State College, PA) can also
process .fsa and .hid files directly as can Cybergenetics’ TrueAllele (Pittsburgh, PA; see also Kadash
2004) and Qualitype’s GenoProof (Dresden, Germany). In addition, the National Center for Biotech-
nology Information (NCBI) has produced an open-source STR genotyping software program (Goor
et al. 2011) called OSIRIS, which stands for Open Source Independent Review and Interpretation
System (OSIRIS 2014).

I. DATA INTERPRETATION
8 1. DATA INTERPRETATION OVERVIEW
D.N.A. BOX 1.1
WHAT INFORMATION IS STORED IN THE .FSA
AND .HID DNA PROFILE COMPUTER FILE?
ABI 310, 3100, 3100-Avant, 3130, 3130xl,
and 3730 Genetic Analyzer instruments (Life
Technologies/Applied Biosystems, Foster City,
CA) produce .fsa files during capillary electro-
phoresis data collection. ABI 3500 and 3500xl in-
struments produce .hid files for human identity
applications and .fsa files for other applications.
During analysis with the Applied Biosystems
software GeneMapperID and GeneMapperID-X
programs, the .fsa or .hid sample files are im-
ported into an Oracle database along with allelic
ladders and other controls for further analysis
(Applied Biosystems 2003, 2004). These Gene-
Mapper projects can be then be exported as .ser
files (Java serialized file) for storage. While .fsa
files can be read by all versions of GeneMapperID
and ID-X, the .hid files can only be read by
GeneMapperID-X v1.2 or above.
The .fsa files are written in a binary file format
known as ABIF (Applied Biosystems, Inc.
Format) and are similar to the TIFF files (Tag
Image File Format) that are sometimes used for
graphics files. Applied Biosystems has published
various versions of their .fsa file format schema,
most recently in September 2009, to enable other
software developers to create products that can
utilize Genetic Analyzer data. Unfortunately, the
full details of their .hid file format have not yet
been publicly released.
However, both .fsa and .hid files appear to
consist of the same basic structure: (1) a header
that points to (2) a directory of tags which then
points via a file offset to (3) electrophoretic data.
The electrophoretic data are collected by scan
number (time) and fluorescence signal in speci-
fied dye-channels. In the developer toolkit on the
Applied Biosystems website, a detailed descrip-
tion of the file tags are provided for (a) ABI 3100
and 3100-Avant, (b) ABI 3130 and 3130xl, (c) ABI
3500 and 3500xl, and (d) ABI 3730 and 3730xl
I. DATA INTERPRETATION
instruments. It appears that the .fsa data file
structure has storage room for up to 99 different
fluorescent dyes, so there is room to grow beyond
the four, five, or even six dyes that STR kit
chemistry currently provide! The .fsa file format
has two sets of tags that point to the same elec-
tropherogram data, while the .hid format consists
of four sets of tags e three essentially in .fsa
format containing the same data, and a fourth set
of tags that are proprietary to Applied Biosystems
software for use in signal normalization. The
fluorescence signal collected by the charged-
coupled device (CCD) camera is stored in a
2-byte format-enabling signal to be collected
between þ32,767 and 32,767. Spectral calibra-
tion, known as “multi-componenting,” is applied
to enable color correction with a mathematical
matrix involving the fluorescent dyes used.
With the introduction of the ABI 3500 and
3500xl Genetic Analyzers in 2010, the .fsa file
format was replaced by .hid files. Initially these
files could only be read by Applied Biosystems
software GeneMapperID-X v1.2 or higher. Now
alternative genotyping software programs such
as GeneMarkerHID (SoftGenetics, State College,
PA), GenoProof (QualiType, Dresden, Germany),
TrueAllele (Cybergenetics, Pittsburgh, PA), and
OSIRIS (National Center for Biotechnology
Information, Bethesda, MD) can process .hid file
formats. The new .hid file format captures addi-
tional information in the sample file including the
3500 radio frequency identification (RFID) infor-
mation used for instrument consumables, such as
the polymer and buffer lot numbers. In addition,
normalization capabilities exist with ABI 3500
.hid files. Normalization, which enables signal to
be equalized between different ABI 3500 or 3500xl
instruments, is performed by multiplying the
data points by a factor calculated from the in-
tensity of some of the internal size standard
Another random document with
no related content on Scribd:
French Beans à la Française 321
(Entremets)
An excellent receipt for French 322
Beans à la Française
To boil Windsor Beans 322
Dressed Cucumbers 322
Mandrang, or Mandram (West 323
Indian receipt)
Another receipt for Mandram 323
Dressed Cucumbers (Author’s 323
receipt)
Stewed Cucumbers (English 323
mode)
Cucumbers à la Poulette 324
Cucumbers à la Créme 324
Fried Cucumbers, to serve in 324
common hashes and minces
Melon 325
To boil Cauliflowers 325
Cauliflowers (French receipt) 325
Cauliflowers with Parmesan 325
Cheese
Cauliflowers à la Française 326
Brocoli 326
To boil Artichokes 326
Artichokes en Salade (see
Chapter VI.)
Vegetable Marrow 327
Roast Tomatas (to serve with 327
roast Mutton)
Stewed Tomatas 327
Forced Tomatas (English 327
receipt)
Forced Tomatas (French 328
receipt)
Purée of Tomatas 328
To boil Green Indian Corn 329
Mushrooms au Beurre 329
Potted Mushrooms 330
Mushroom-Toast, or Croule 330
aux Champignons (excellent)
Truffles, and their uses 331
Truffles à la Serviette 331
Truffles à l’Italienne 331
To prepare Truffles for use 332
To boil Sprouts, Cabbages, 332
Savoys, Lettuces, or Endive
Stewed Cabbage 333
To boil Turnips 333
To mash Turnips 333
Turnips in white Sauce 334
(Entremets)
Turnips stewed in Butter (good) 334
Turnips in Gravy 335
To boil Carrots 335
Carrots (the Windsor receipt) 335
(Entremets)
Sweet Carrots (Entremets) 336
Mashed (or Buttered) Carrots 336
(a Dutch receipt)
Carrots au Beurre, or Buttered 336
Carrots (French receipt)
Carrots in their own Juice (a 337
simple but excellent receipt)
To boil Parsneps 337
Fried Parsneps 337
Jerusalem Artichokes 337
To fry Jerusalem Artichokes 338
(Entremets)
Jerusalem Artichokes à la 338
Reine
Mashed Jerusalem Artichokes 338
Haricots Blancs 338
To boil Beet-Root 339
To bake Beet-Root 339
Stewed Beet-Root 340
To stew Red Cabbage (Flemish 340
receipt)
Brussels Sprouts 340
Salsify 341
Fried Salsify (Entremets) 341
Boiled Celery 341
Stewed Celery 341
Stewed Onions 342
Stewed Chestnuts 342
CHAPTER XVIII.
 PASTRY.

Page

Introductory remarks 344

To glaze or ice Pastry 345
Feuilletage, or fine French Puff 345
Paste
Very good light Paste 346
English Puff Paste 346
Cream Crust (very good) 347
(Author’s receipt)
Pâte Brisée (or French Crust 347
for hot or cold Meat Pies)
Flead Crust 347
Common Suet-Crust for Pies 348
Very superior Suet-Crust 348
Very rich short Crust for Tarts 349
Excellent short Crust for Sweet 349
Pastry
Bricche Paste 349
Modern Potato Pasty, an 350
excellent family dish
Casserole of Rice 351
A good common English Game 352
Pie
Modern Chicken Pie 353
A common Chicken Pie 353
Pigeon Pie 354
Beef-steak Pie 354
Common Mutton Pie 355
A good Mutton Pie 355
Raised Pies 356
A Vol-au-Vent (Entrée) 357
A Vol-au-Vent of Fruit 358
(Entremets)
A Vol-au-Vent à la Créme 358
(Entremets)
Oyster Patties (Entrée) 359
Common Lobster Patties 359
Superlative Lobster Patties 359
(Author’s receipt)
Good Chicken Patties (Entrée) 359
Patties à la Pontife, a fast-day 360
or maigre dish (Entrée)
Excellent Meat Rolls 360
Small Vols-au-Vents, or Patty- 361
cases
Another receipt for Tartlets 361
A Sefton, or Veal Custard 362
Apple Cake, or German Tart 362
Tourte Meringuée, or Tart with 363
royal icing
A good Apple Tart 363
Tart of very young green 364
Apples (good)
Barberry Tart 364
The Lady’s Tourte, and 364
Christmas Tourte à la
Châtelaine
Genoises à la Reine, or her 366
Majesty’s Pastry
Almond Paste 367
Tartlets of Almond Paste 367
Fairy Fancies (Fantaisies des 368
Fées)
Mincemeat (Author’s receipt) 368
Superlative Mincemeat 369
Mince Pies (Entremets) 369
Mince Pies Royal (Entremets) 370
The Monitor’s Tart, or Tourte à 370
la Judd
Pudding Pies (Entremets) 371
Pudding Pies (a commoner 371
kind)
Cocoa-Nut cheese-cakes 371
(Entremets) (Jamaica
receipt)
Common Lemon Tartlets 372
Madame Werner’s Rosenvik 372
cheese-cakes
Apfel Krapfen (German receipt) 373
Créme Pâtissière, or Pastry 373
Cream
Small Vols-au-Vent, à la 374
Parisienne (Entremets)
Pastry Sandwiches 374
Lemon Sandwiches 374
Fanchonnettes (Entremets) 374
Jelly-Tartlets, or Custards 375
Strawberry Tartlets (good) 375
Raspberry Puffs 375
Creamed Tartlets 375
Ramakins à l’Ude, or Sefton- 375
Fancies
 CHAPTER XIX.

SOUFFLÉS, OMLETS, ETC.

Page

Soufflés 377
Louise Franks’ Citron Soufflé 378
A Fondu, or Cheese Souffle 379
Observations on Omlets, 380
Fritters, &c.
A common Omlet 380
An Omlette Soufflé (second 381
course, remove of roast)
Plain Common Fritters 381
Pancakes 382
Fritters of Cake and Pudding 382
Mincemeat Fritters 383
Venetian Fritters (very good) 383
Rhubarb Fritters 383
Apple, Peach, Apricot, or 384
Orange Fritters
Brioche Fritters 384
Potato Fritters (Entremets) 384
Lemon Fritters (Entremets) 384
Cannelons (Entremets) 385
Cannelons of Brioche paste 385
(Entremets)
Croquettes of Rice (Entremets) 385
Finer Croquettes of Rice 386
(Entremets)
Savoury Croquettes of Rice 386
(Entrée)
Rissoles (Entrée) 387
Very savoury Rissoles (Entrée) 387
Small fried Bread Patties, or 387
Croustades of various kinds
Dresden Patties, or Croustades 387
(very delicate)
To prepare Beef Marrow for 388
frying Croustades, Savoury
Toasts, &c.
Small Croustades, or Bread 388
Patties, dressed in Marrow
(Author’s receipt)
Small Croustades, à la Bonne 389
Maman (the Grandmamma’s
Patties)
Curried Toasts with Anchovies 389
To fillet Anchovies 389
Savoury Toasts 390
To choose Macaroni, and other 390
Italian Pastes
To boil Macaroni 391
Ribbon Macaroni 391
Dressed Macaroni 392
Macaroni à la Reine 393
Semoulina and Polenta à 393
l’Italienne (Good) (To serve
instead of Macaroni)
 CHAPTER XX.

BOILED PUDDINGS.

Page

General Directions 395

To clean Currants for Puddings 397
or Cakes
To steam a Pudding in a 397
common stewpan or
saucepan
To mix Batter for Puddings 397
Suet Crust for Meat or Fruit 398
Pudding
Butter Crust for Puddings 398
Savoury Puddings 399
Beef-steak, or John Bull’s 399
Pudding
Small Beef-steak Pudding 400
Ruth Pinch’s Beef-steak 401
Pudding
Mutton Pudding 401
Partridge Pudding (very good) 401
A Peas Pudding (to serve with 401
Boiled Pork)
Wine-sauce for Sweet 402
Puddings
Common Wine-sauce 402
Punch-sauce for Sweet 402
Puddings
Clear arrow-root-sauce (with 403
receipt for Welcome Guest’s
Pudding)
A German Custard Pudding- 403
sauce
A delicious German Pudding- 403
sauce
Red Currant or Raspberry- 404
sauce (good)
Common Raspberry-sauce 404
Superior Fruit Sauces for 404
Sweet Puddings
Pine-apple Pudding-sauce 405
A very fine Pine-apple Sauce 405
or Syrup for Puddings, or
other Sweet Dishes
German Cherry-sauce 406
Common Batter Pudding 406
Another Batter Pudding 406
Black-cap Pudding 407
Batter Fruit Pudding 407
Kentish Suet Pudding 407
Another Suet Pudding 408
Apple, Currant, Cherry, or other 408
Fresh Fruit Pudding
A common Apple Pudding 409
Herodotus’ Pudding (A genuine 409
classical receipt)
The Publisher’s Pudding 410
Her Majesty’s Pudding 410
Common Custard Pudding 411
Prince Albert’s Pudding 411
German Pudding and Sauce 412
(very good)
The Welcome Guest’s own 412
Pudding (light and
wholesome. Author’s receipt)
Sir Edwin Landseer’s Pudding 412
A Cabinet Pudding 413
A very fine Cabinet Pudding 414
Snowdon Pudding (a genuine 414
receipt)
Very good Raisin Puddings 415
The Elegant Economist’s 415
Pudding
Pudding à la Scoones 416
Ingoldsby Christmas Puddings 416
Small and very light Plum 416
Pudding
Vegetable Plum Pudding 417
(cheap and good)
The Author’s Christmas 417
Pudding
A Kentish Well-Pudding 417
Rolled Pudding 418
A Bread Pudding 418
A Brown Bread Pudding 419
A good boiled Rice Pudding 419
Cheap Rice Pudding 420
Rice and Gooseberry Pudding 420
Fashionable Apple Dumplings 420
Orange Snow-balls 420
Apple Snow-balls 421
Light Currant Dumplings 421
Lemon Dumplings (light and 421
good)
Suffolk, or hard Dumplings 421
Norfolk Dumplings 421
Sweet boiled Patties (good) 422
Boiled Rice, to be served with 422
stewed Fruits, Preserves, or
Raspberry Vinegar
 CHAPTER XXI.

BAKED PUDDINGS.

Page

Introductory Remarks 423

A baked Plum Pudding en 424
Moule, or Moulded
The Printer’s Pudding 424
Almond Pudding 425
The Young Wife’s Pudding 425
(Author’s receipt)
The Good Daughter’s 426
Mincemeat Pudding
(Author’s receipt)
Mrs. Howitt’s Pudding (Author’s 426
receipt)
An excellent Lemon Pudding 426
Lemon Suet Pudding 427
Bakewell Pudding 427
Ratifia Pudding 427
The elegant Economist’s 428
Pudding
Rich Bread and Butter Pudding 428
A common Bread and Butter 429
Pudding
A good baked Bread Pudding 429
Another baked Bread Pudding 430
A good Semoulina or Soujee 430
Pudding
French Semoulina Pudding, or 430
Gâteau de Semoule
Saxe-Gotha Pudding, or Tourte 431
Baden Baden Puddings 431
Sutherland, or Castle Puddings 432
Madeleine Puddings (to be 432
served cold)
A good French Rice Pudding, 433
or Gâteau de Riz
A common Rice Pudding 433
Quite cheap Rice Pudding 434
Richer Rice Pudding 434
Rich Pudding Meringué 434
Good ground Rice Pudding 435
Common ground Rice Pudding 435
Green Gooseberry Pudding 435
Potato Pudding 436
A Richer Potato Pudding 436
A good Sponge-cake Pudding 436
Cake and Custard, and various 437
other inexpensive Puddings
Baked Apple Pudding, or 437
Custard
Dutch Custard, or Baked 438
Raspberry Pudding
Gabrielle’s Pudding, or sweet 438
Casserole of Rice
Vermicelli Pudding, with apples 439
or without, and Puddings of
Soujee and Semola
Rice à la Vathek, or Rice 440
 Pudding à la Vathek
(extremely good)
Good Yorkshire Pudding 440
Common Yorkshire Pudding 441
Normandy Pudding (good) 441
Common baked Raisin 441
Pudding
A richer baked Raisin Pudding 442
The Poor Author’s Pudding 442
Pudding à la Paysanne (cheap 442
and good)
The Curate’s Pudding 442
A light baked Batter Pudding 443
 CHAPTER XXII.

EGGS AND MILK.

Page

To preserve Eggs fresh for 444

many weeks
To cook Eggs in the shell 445
without boiling them (an
admirable receipt)
To boil Eggs in the shell 445
To dress the Eggs of the 446
Guinea Fowl and Bantam
To dress Turkeys’ Eggs 447
Forced Turkeys’ Eggs (or 447
Swans’), an excellent
entremets
To boil a Swan’s Egg hard 448
Swan’s Egg en Salade 448
To poach Eggs of different 449
kinds
Poached Eggs with Gravy 449
(Œufs Pochés au Jus.
Entremets.)
Œufs au Plat 450
Milk and Cream 450
Devonshire, or Clotted Cream 451
Du Lait a Madame 451
Curds and Whey 451
Devonshire Junket 452
 CHAPTER XXIII.

SWEET DISHES, OR ENTREMETS.

Page

To prepare Calf’s Feet Stock 453

To clarify Calf’s Feet Stock 454
To clarify Isinglass 454
Spinach Green, for colouring 455
Sweet Dishes,
Confectionary, or Soups
Prepared Apple or Quince 456
Juice
Cocoa-nut flavoured Milk (for 456
Sweet Dishes, &c.)
Remarks upon Compotes of 456
Fruit, or Fruit stewed in
Syrup
Compote of Rhubarb 457
—— of Green Currants 457
—— of Green Gooseberries 457
—— of Green Apricots 457
—— of Red Currants 457
—— of Raspberries 458
—— of Kentish or Flemish 458
Cherries
—— of Morella Cherries 458
—— of the green Magnum 458
Bonum, or Mogul Plum
—— of Damsons 458
—— of ripe Magnum Bonums, 458
or Mogul Plums
—— of the Shepherd’s and 458
other Bullaces
—— of Siberian Crabs 458
—— of Peaches 459
Another receipt for stewed 459
Peaches
Compote of Barberries for 459
Dessert
Black Caps, par excellence (for 460
the Second Course, or for
Dessert)
Gâteau de Pommes 460
Gâteau of mixed Fruits (good) 461
Calf’s Feet Jelly (entremets) 461
Another receipt for Calf’s Feet 462
Jelly
Modern varieties of Calf’s Feet 463
Jelly
Apple Calf’s Feet Jelly 464
Orange Calf’s Feet Jelly 464
(Author’s receipt)
Orange Isinglass Jelly 465
Very fine Orange Jelly (Sussex 465
Place receipt)
Oranges filled with Jelly 466
Lemon Calf’s Feet Jelly 467
Constantia Jelly 467
Rhubarb Isinglass Jelly 468
(Author’s original receipt)