Genomics Protocols

538 Pages·2001·3.512 MB·English

by  Paul A. Lyons (auth.), Michael P. Starkey, Ramnath Elaswarapu (eds.)

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Genomics Protocols
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M E T H O D S I N M O L E C U L A R B I O L O G Y™
Genomics Protocols
Edited by
Michael P. Starkey
and
Ramnath Elaswarapu
UK Human Genome Mapping Project Resource Centre,
Cambridge, UK
Humana Press Totowa, New Jersey
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Genomics protocols / edited by Michael P. Starkey and Ramnath Elaswarapu.
p. ; cm.—(Methods in molecular biology ; 175)
Includes bibliographical references and index.
ISBN 0-89603-774-6 (hardcover ; alk. paper)--ISBN 0-89603-708-8 (comb ; alk. paper)
1. Molecular genetics—Laboratory manuals. 2. Genomics–Laboratory manuals. I. Starkey, Michael P. II.
Elaswarapu, Ramnath. III. Series QH440.5 G46 2001]
572.8—dc21
00-046106
Preface
We must unashamedly admit that a large part of the motivation for editing
Genomics Protocols was selfish. The possibility of assembling in a single volume
a unique and comprehensive collection of complete protocols, relevant to our
work and the work of our colleagues, was too good an opportunity to miss.
We are pleased to report, however, that the outcome is something of use not
only to those who are experienced practitioners in the genomics field, but is
also valuable to the larger community of researchers who have recognized the
potential of genomics research and may themselves be beginning to explore
the technologies involved.
Some of the techniques described in Genomics Protocols are clearly not
restricted to the genomics field; indeed, a prerequisite for many procedures in
this discipline is that they require an extremely high throughput, beyond the
scope of the average investigator. However, what we have endeavored here to
achieve is both to compile a collection of procedures concerned with genome-
scale investigations and to incorporate the key components of “bottom-up”
and“top-down” approaches to gene finding. The technologies described extend
from those traditionally recognized as coming under the genomics umbrella,
touch on proteomics (the study of the expressed protein complement of the
genome), through to early therapeutic approaches utilizing the potential of
genome programs via gene therapy (Chapters 27–30).
Although a number of the procedures described represent the tried and
trusted, we have striven to include new variants on existing technologies in
addition to exciting new approaches. Where there are alternative approaches
to achieving a particular goal, we have sought assistance from an expert in the
field to identify the most reliable technique, one suitable for a beginner in the
field. Unique to the Methods in Molecular Biology series is the “Notes” section at
the end of each chapter. This is a veritable Aladdin’s cave of information in
which an investigator describes the quirks in a procedure and the little tricks
that make all the difference to a successful outcome.
The first section of the volume deals with the traditional positional cloning
approach to gene identification and isolation. The construction of a high-reso-
lution genetic map (Chapter 1) to facilitate the mapping of monogenic traits
v
vi Preface
and approaches to the analysis of polygenic traits (Chapter 2) are described.
Identification of large numbers of single-nucleotide polymorphisms (Chapter
3) will pave the way for the construction of the next generation of genetic
maps. Also described are such comparatively new technologies as genomic
mismatch scanning (Chapter 4), for the mapping of genetic traits, and compar-
ative genomic hybridization (Chapter 5), for the identification of gross differ-
ences between genomes.
Such studies are a prelude to the screening of large genomic clones, or
clone contigs (Chapter 7). These transitions are made possible by the locali-
zation of genomic clones (Chapter 8) and the integration of the genetic and
physical maps (Chapter 9) achieved by STS mapping. Identification of cDNAs
mapping to the genomic clones implicated (Chapters 12–14) is the next step
toward candidate gene identification. With the desire to acquire cDNAs capable
of expressing authentic proteins, the emphasis in cDNA library construction
is placed on a technology capable of delivering full-length cDNAs (Chapter 10).
One of the consequences of genome-scale sequencing programs has been
the need to annotate large stretches of anonymous sequence data, and this has
been the impetus for an explosion of bioinformatics programs targeted at gene
prediction (Chapter 16). The use of model organisms (Chapter 17) to expedite
gene discovery, on the basis of coding sequence similarites between genes
with similiar functions, is another tool accessible to the gene hunter.
As an alternative to genetic studies, expression profiling seeks to iden-
tify candidate genes on the basis of their differential patterns of expression,
either at the level of transcription or translation. A number of technologies,
based on subtractive hybridization, differential display, and high throughput
in situ hybridization are thus described (Chapters 18–22).
Functional characterization of isolated cDNAs is the next stage in estab-
lishing the likely candidature and thus potential utility of genes isolated as
targets for therapeutic intervention. Predictions of protein structure and function
(Chapter 23), mutagenesis (Chapter 24), or knockout studies (Chapter 25) can
enable predictions of gene function. The yeast two-hybrid system (Chapter 26)
is described at the level of monitoring interaction between individual proteins,
but also on a potential genome scale.
In compiling Genomics Protocols, the aim—as with all other volumes in
the Methods in Molecular Biology series—has been to produce a self-contained
laboratory manual useful to both experienced practitioners and beginners in
the field. We trust that we have been at least moderately successful. We must
conclude by giving a vote of thanks to all the contributing authors, and to John
Walker and the staff at Humana Press for seeing this project through.
Michael P. Starkey
Ramnath Elaswarapu
Contents
Preface .................................................................................................v
Contributors.........................................................................................xi
1 Construction of Microsatellite-Based, High-Resolution
Genetic Maps in the Mouse
Paul A. Lyons................................................................................1
2 Genetic Analysis of Complex Traits
Stephen P. Bryant and Mathias N. Chiano..............................11
3 Sequence-Based Detection
of Single Nucleotide Polymorphisms
Deborah A. Nickerson, Natali Kolker, Scott L. Taylor,
and Mark J. Rieder................................................................29
4 Genomic Mismatch Scanning for the Mapping of Genetic Traits
Farideh Mirzayans and Michael A. Walter...............................37
5 Detection of Chromosomal Abnormalities by Comparative
Genomic Hybridization
Mario A. J. A. Hermsen, Marjan M. Weiss, Gerrit A. Meijer,
and Jan P. A. Baak...............................................................47
6 Construction of a Bacterial Artificial Chromosome Library
Sangdun Choi and Ung-Jin Kim...............................................57
7 Contiguation of Bacterial Clones
Sean J. Humphray, Susan J. Knaggs,
and Ioannis Ragoussis........................................................69
8 Mapping of Genomic Clones by Fluorescence In Situ
Hybridization
Margaret A. Leversha...............................................................109
9 Map Integration: From a Genetic Map to a Physical Gene Map
and Ultimately to the Sequence Map
Panagiotis Deloukas................................................................129
10 Construction of Full-Length–Enriched cDNA Libraries:
The Oligo-Capping Method
Yutaka Suzuki and Sumio Sugano.........................................143
vii
viii Contents
11 Construction of Transcript Maps by Somatic Cell/Radiation
Hybrid Mapping: The Human Gene Map
Panagiotis Deloukas................................................................155
12 Preparation and Screening of High-Density cDNA Arrays
with Genomic Clones
Günther Zehetner, Maria Pack, and Katja Schäfer...............169
13 Direct Selection of cDNAs by Genomic Clones
Daniela Toniolo.........................................................................189
14 Exon Trapping: Application of a Large-Insert
Multiple-Exon-Trapping System
Martin C. Wapenaar and Johan T. Den Dunnen...................201
15 Sequencing Bacterial Artificial Chromosomes
David E. Harris and Lee Murphy.............................................217
16 Finding Genes in Genomic Nucleotide Sequences
by Using Bioinformatics
Yvonne J. K. Edwards and Simon M. Brocklehurst.............235
17 Gene Identification Using the Pufferfish, Fugu rubripes,
by Sequence Scanning
Greg Elgar.................................................................................249
18 Isolation of Differentially Expressed Genes
Through Subtractive Suppression Hybridization
Oliver Dorian von Stein...........................................................263
19 Isolation of Differentially Expressed Genes
by Representational Difference Analysis
Christine Wallrapp and Thomas M. Gress............................279
20 Expression Profiling and the Isolation of Differentially
Expressed Genes by Indexing-Based Differential Display
Michael P. Starkey....................................................................295
21 Expression Profiling by Systematic High-ThroughputIn Situ
Hybridization to Whole-Mount Embryos
Nicolas Pollet and Christof Niehrs.........................................309
22 Expression Monitoring Using cDNA Microarrays:
A General Protocol
Xing Jian Lou, Mark Schena, Frank T. Horrigan,
Richard M. Lawn, and Ronald W. Davis..........................323
23 Prediction of Protein Structure and Function
by Using Bioinformatics
Yvonne J. K. Edwards and Amanda Cottage........................341
Contents ix
24 Identification of Novel Genes by Gene Trap Mutagenesis
Anne K. Voss and Tim Thomas..............................................377
25 Determination of Gene Function by Homologous Recombination
Using Embryonic Stem Cells and Knockout Mice
Ahmed Mansouri......................................................................397
26 Genomic Analysis Utilizing the Yeast Two-Hybrid System
Ilya G. Serebriiskii, Garabet G. Toby, Russell L. FInley, Jr.,
and Erica A. Golemis...........................................................415
27 Methods for Adeno-Associated Virus–Mediated
Gene Transfer into Muscle
Terry J. Amiss and Richard Jude Samulski..........................455
28 Retroviral-Mediated Gene Transduction
Donald S. Anson.......................................................................471
29 Gene Therapy Approaches to Sensitization of Human Prostate
Carcinoma to Cisplatin by Adenoviral Expression of p53
and by Antisense Jun Kinase Oligonucleotide Methods
Ruth Gjerset, Ali Haghighi, Svetlana Lebedeva,
and Dan Mercola.................................................................495
30 Ribozyme Gene Therapy
Leonidas A. Phylactou.............................................................521
Index.................................................................................................531