Analytical Techniques in DNA Sequencing - B. Nunnally (CRC, 2005) WW

250 Pages·2005·4.51 MB·English

ANALYTICAL
TECHNIQUES
in DNA
SEQUENCING

DK1883_C000.fm Page ii Saturday, January 16, 1904 12:58 AM

ANALYTICAL
TECHNIQUES
in DNA
SEQUENCING

E d i t e d b y
B r i a n K . N u n n a l l y

Boca Raton London New York Singapore

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International Standard Book Number-10: 0-8247-5342-9 (Hardcover)
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Library of Congress Cataloging-in-Publication Data

Analytical techniques in DNA sequencing / edited by Brian K. Nunnally.
p. cm.
Includes bibliographical references and index.
ISBN 0-8247-5342-9 (alk. paper)
1. Nucleotide sequence. I. Nunnally, Brian K.

QP625.N89A53 2005
611'.01816--dc22
2004066429

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Preface

The modern era of DNA sequencing began in late 1977 with the introduction of the
most common DNA sequencing method, the Sanger method. The Sanger method
involves the use of radioactive dideoxynucleotides, a deoxynucleotide with the 3′
hydroxyl group from the deoxyribose sugar removed.1 The Sanger method relies on
statistics to create fragments that are terminated at every position of the DNA. The
presence of a band indicates the base position and identity.
Fluorescence-based sequencing was introduced in 1986 by L.M. Smith, L.E. Hood,
and coworkers.2 Four different fluorescent dyes were attached to the dideoxynucle-
otides allowing for spectral discrimination of the fragments. Fluorescence shows com-
parable performance and is an attractive alternative due to its relative safety, real-time
capability, and ease of automation, as well as the ability to multiplex. In addition, the
fluorescent dyes do not significantly affect the fidelity of the enzymes.3

The first multiplex fluorescence-based sequencing systems used a four-channel
approach, similar to the radioactive-based sequencing.4 Smith, Hood, and coworkers2
used a set of four dyes with different emission maxima. The signal was selected
using different interference filters based on the different dye emission maxima.
The first system of dyes included the following: fluorescein isothiocyanate
(λem = 516 nm), NBD-aminohexanoic acid (λem = 540 nm), tetramethylrhodamine
isothiocyanate (λem = 582 nm), and Texas Red (λem = 612 nm). This system was
later commercialized by Applied Biosystems (ABI) using a revised set of dyes: FAM
(λem = 521 nm), JOE (λem = 555 nm), TAMRA (λem = 580 nm), and ROX
(λem = 605 nm).5 This commercial system allowed analytical sequencing to become
a popular and routine technique for many laboratories. Mobility corrections are
needed for this dye system. Ju et al.6 developed a novel energy transfer system that
addressed some of the failings of this system, which allowed for more efficient
excitation with equal mobilities.
The original DNA sequencing systems were based on the standard slab polyacryl-
amide gel electrophoresis equipment, which allowed numerous samples to be analyzed
on the same gel. Not long after the introduction of the slab-gel sequencing systems,
a capillary electrophoresis (CE)-based sequencing system was developed. The CE
system permitted increased speed, ease of use, and increased accuracy, although the
CE system had a much lower throughput than the slab-gel system until the development
of multicapillary systems. These systems are now commercially available and use from
8 to 96 capillaries in large arrays. Other techniques such as MALDI MS have been
tried with modest success, but have no significant application. The future of DNA
sequencing may lie in the use of microfabricated sequencing systems. These chip-
based techniques will allow DNA sequencing to expand into a variety of new envi-
ronments.
From the early days of radioisotope sequencing, a wide variety of new techniques
have emerged to meet the needs of biotechnology. Techniques aimed at reducing the

amount of sample needed, improving the accuracy, and reducing the amount of time
needed to generate a sequence have been employed. The focus of this book is discussion
of the different analytical DNA sequencing techniques, as well as some of the exciting
applications of DNA sequencing. Numerous applications are discussed in this book,
including microbiological identification, forensic DNA sequencing, and ancient DNA
sequencing.

REFERENCES

1. F Sanger, S Nicklen, AR Coulson. Proc Natl Acad Sci USA 74, 5463–5467, 1977.
2. LM Smith, JZ Sander, RJ Kaiser, P Hughes, C Dodd, CR Connell, C Heiner, SBH
Kent, LE Hood. Nature 321, 674–679, 1986.
3. LM Smith, S Fung, MW Hunkapiller, TJ Hunkapiller, LE Hood. Nucleic Acids Res
13, 2399–2412, 1985.
4. H Swerdlow, JZ Zhang, DY Chen, HR Harke, R Grey, S Wu, NJ Dovichi, C Fuller.
Anal Chem 63, 2385–2841, 1991.
5. C Connell, S Fung, C Heiner, J Bridgham, V Chakerian, E Heron, B Jones,
S Menchen, W Mordan, M Raff, M Recknor, L Smith, J Springer, S Woo,
M Hunkapiller. BioTechniques 5, 342–348, 1987.
6. J Ju, AN Glazer, RA Mathies. Nat Med 2, 246–249, 1996.

Editor

Brian K. Nunnally, Sr., Ph.D., received his Ph.D. in chemistry and a certificate in
molecular biophysics from Duke University, Durham, NC, in 1998. He is assistant
director at Wyeth Laboratories in Sanford, NC, where he works in Vaccine Analytical
Development. Dr. Nunnally’s research interests lie in bioanalytical chemistry, includ-
ing research on protein and polysaccharide therapeutics and BSE/TSE issues (BSE,
bovine spongiform encephalopathy; TSE, transmissible spongiform encephalopa-
thy). Prior to joining Wyeth, he worked for Eli Lilly and Company as a research
scientist in quality control.
Dr. Nunnally has published numerous articles and lectured on a variety of analyt-
ical and pharmaceutical disciplines. His graduate work focused on the development
of multiplex detection for bioanalytical separations using fluorescence lifetime. This
included DNA sequencing fragment separations. In 2004, the book Dr. Nunnally
coedited with Professor Ira Krull, titled Prions and Mad Cow Disease (Marcel Dekker,
New York), was published.
Dr. Nunnally currently serves as assistant editor for Analytical Letters (CRC Press,
Boca Raton, FL). He served as the cochair of the CE in the Biotechnology and
Pharmaceutical Industries Conference in 2003 and 2004. Dr. Nunnally is a member
of the Analytical Chemistry Division of the American Chemical Society and of the
Society for Applied Spectroscopy. He was graduated with two honors degrees from
University of South Carolina, Columbia. Dr. Nunnally is an Eagle Scout and father
of two children, Brian Jr. and Annabelle.

Contributors

Robert G. Blazej
Department of Chemistry
and UCSF/UCB Joint Graduate Group
in Bioengineering
University of California
Berkeley, California

William Goodwin
Department of Forensic Medicine
and Science
University of Glasgow
Glasgow, Scotland

Samuel A. Heath
Computer Science Department
Brown University
Providence, Rhode Island

Franz Hillenkamp
Institute for Medical Physics
and Biophysics
University of Münster
Münster, Germany

Dorrie Main
Clemson University Genomics Institute
Clemson, South Carolina

Richard A. Mathies
Department of Chemistry
University of California
Berkeley, California

Terry Melton
Mitotyping Technologies, LLC
State College, Pennsylvania

Brian K. Nunnally
Wyeth Laboratories
Sanford, North Carolina

Brian M. Paegel
Department of Chemistry
University of California
Berkeley, California

Franco P. Preparata
Computer Science Department
Brown University
Providence, Rhode Island

Markus Sauer
Applied Laserphysics and
Laserspectroscopy
University of Bielefeld
Bielefeld, Germany

Jeffrey P. Tomkins
Clemson University Genomics Institute
Clemson, South Carolina

Eli Upfal
Computer Science Department
Brown University
Providence, Rhode Island

Dirk van den Boom
Sequenom, Inc.
San Diego, California

Victor W. Weedn
Carnegie Mellon University
Mellon Institute
Pittsburgh, Pennsylvania