DNA Replication: Methods and Protocols

298 Pages·2015·7.194 MB·English

by  Sonya Vengrova, Jacob Dalgaard (eds.)

Methods in
Molecular Biology 1300
Sonya Vengrova
Jacob Dalgaard Editors
DNA
Replication
Methods and Protocols
Second Edition
M M B
ETHODS IN OLECULAR IOLOGY
Series Editor
John M. Walker
School of Life and Medical Sciences
University of Hertfordshire
Hat fi eld, Hertfordshire, AL10 9AB, UK
For further volumes:
http://www.springer.com/series/7651

DNA Replication
Methods and Protocols
Second Edition
Edited by
Sonya Vengrova
Coventry, UK
Jacob Dalgaard
Warwick Medical School, University of Warwick, Coventry, UK
Editors
Sonya Vengrova Jacob D algaard
Coventry, UK Warwick Medical School
University of Warwick
Coventry, UK
ISSN 1064-3745 ISSN 1940-6029 (electronic)
Methods in Molecular Biology
ISBN 978-1-4939-2595-7 ISBN 978-1-4939-2596-4 (eBook)
DOI 10.1007/978-1-4939-2596-4
Library of Congress Control Number: 2015935079
Springer New York Heidelberg Dordrecht London
© Springer Science+Business Media New York 2 015
This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is
concerned, specifi cally the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction
on microfi lms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation,
computer software, or by similar or dissimilar methodology now known or hereafter developed.
The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not
imply, even in the absence of a specifi c statement, that such names are exempt from the relevant protective laws and
regulations and therefore free for general use.
The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to
be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty,
express or implied, with respect to the material contained herein or for any errors or omissions that may have been made.
Printed on acid-free paper
Humana Press is a brand of Springer
Springer Science+Business Media LLC New York is part of Springer Science+Business Media (www.springer.com)
Prefa ce
The research fi eld of DNA replication is progressing rapidly, and new technologies are
developed every year, allowing researches to understand novel aspects of the complex
processes of DNA replication. In this volume of “DNA Replication: Methods and Protocols”
17 method chapters are presented, which cover a range of systems, including mammalian,
yeast, bacterial, and archaeal. The methods describe approaches to analyze whole genomes
to single molecules, as well as both in vivo and in vitro experiments. This collection con-
tains highly sited updated versions of methods fi rst published in the previous edition as well
as novel methods that have been developed in the recent years.
The methods are described in a way that enables researchers new to the fi eld to execute
them; some of them can be performed using basic molecular biology equipment, while oth-
ers require specialized setup.
As a whole, this method collection is intended for both newcomers to this research fi eld
and established laboratories.
Coventry, UK Sonya Vengrova
J acob D algaard
v

Contents
Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . i x
1 A High-Throughput Confocal Fluorescence Microscopy Platform
to Study DNA Replication Stress in Yeast Cells . . . . . . . . . . . . . . . . . . . . . . . . 1
Nikko P . Torres and Grant W . B rown
2 M icroscopy Techniques to Examine DNA Replication
in Fission Yeast . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 3
Marc D. Green , Sarah A . Sabatinos , and Susan L. Forsburg
3 H igh-Resolution Analysis of Mammalian DNA Replication Units . . . . . . . . . . 4 3
Vadim O. C hagin , M arius Reinhart , and M . C ristina C ardoso
4 A nalyzing the Dynamics of DNA Replication in Mammalian Cells
Using DNA Combing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67
Marta B ialic , V incent C oulon , Marjorie D rac , T hierry G ostan ,
and Etienne Schwob
5 M easuring DNA Content by Flow Cytometry in Fission Yeast. . . . . . . . . . . . . 7 9
Sarah A . Sabatinos and Susan L . F orsburg
6 I ncorporation of Thymidine Analogs for Studying Replication
Kinetics in Fission Yeast. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 99
Nicholas R hind
7 E dU Incorporation for FACS and Microscopy Analysis
of DNA Replication in Budding Yeast. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105
Nicolas T alarek , Julie Petit , E lisabeth G ueydon , and Etienne Schwob
8 D etermination of Deoxyribonucleoside Triphosphate Concentrations
in Yeast Cells by Strong Anion-Exchange High-Performance
Liquid Chromatography Coupled with Ultraviolet Detection . . . . . . . . . . . . . 1 13
Shaodong J ia , Lisette M arjavaara , Robert B uckland , S ushma S harma ,
and A ndrei Chabes
9 M easuring Ribonucleotide Incorporation into DNA In Vitro and In Vivo. . . . 1 23
Anders R . Clausen , Jessica S . W illiams , and Thomas A . Kunkel
10 D etection and Sequencing of Okazaki Fragments in S. cerevisiae . . . . . . . . . . . 1 41
Duncan J. S mith , Tejas Y adav , and Iestyn Whitehouse
11 ChIP-Seq to Analyze the Binding of Replication Proteins to Chromatin . . . . . 1 55
A. Z achary O strow , Christopher J . V iggiani , Jennifer G. A paricio ,
and Oscar M. Aparicio
12 C hromatin Immunoprecipitation to Detect DNA Replication
and Repair Factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 69
Mariana C . G adaleta , Osamu I wasaki , Chiaki N oguchi ,
Ken-Ichi Noma , and Eishi N oguchi
vii
viii Contents
13 Molecular Genetic Methods to Study DNA Replication Protein Function
in Haloferax volcanii, A Model Archaeal Organism . . . . . . . . . . . . . . . . . . . . . 187
Xavier Giroux and S tuart A . M acNeill
14 S ingle-Molecule Observation of Prokaryotic DNA Replication . . . . . . . . . . . . 2 19
Hylkje J. Geertsema , Karl E. Duderstadt , and Antoine M. van Oijen
15 Analyzing the Response to Dysfunction Replication Forks
Using the RTS1 Barrier System in Fission Yeast. . . . . . . . . . . . . . . . . . . . . . . . 239
Saed M ohebi , S arah A . E. Lambert , and Antony M . C arr
16 Purification of Restriction Fragments Containing Replication
Intermediates from Complex Genomes for 2-D Gel Analysis. . . . . . . . . . . . . . 261
Larry D. Mesner , Pieter A. D ijkwel , and Joyce L. Hamlin
17 I solation of Restriction Fragments Containing Origins
of Replication from Complex Genomes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 79

Larry D. Mesner and J oyce L . H amlin
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 93
Contributors
JENNIFER G. APARICIO • Division of Ophthalmology , C hildren’s Hospital Los Angeles ,
Los Angeles, C A , U SA
OSCAR M. APARICIO • Molecular and Computational Biology Program , U niversity
of Southern California , L os Angeles, CA , U SA
MARTA BIALIC • Institute of Molecular Genetics, CNRS UMR5535 & Université Montpellier ,
Montpellier, F rance
GRANT W . BROWN • Department of Biochemistry and Donnelly Centre , U niversity
of Toronto , T oronto , O N , Canada
ROBERT B UCKLAND • Department of Medical Biochemistry and Biophysics , U mea University ,
Umea, S weden
M. C RISTINA CARDOSO • Department of Biology, Cell Biology and Epigenetics, T echnische
Universität Darmstadt , Darmstadt, Germany
ANTONY M . CARR • MRC Genome Damage and Stability Centre, School of Life Sciences ,
University of Sussex , Brighton, UK
ANDREI CHABES • Department of Medical Biochemistry and Biophysics , U mea University ,
Umea , S weden
VADIM O . CHAGIN • Institute of Cytology, Russian Academy of Science , S aint Petersburg,
Russia
ANDERS R . CLAUSEN • Genome Integrity and Structural Biology Laboratory , N ational
Institute of Environmental Health Sciences, NIH , R esearch Triangle Park, NC, U SA
VINCENT COULON • Institute of Molecular Genetics, CNRS UMR5535 & Université
Montpellier , M ontpellier, France
PIETER A. DIJKWEL • Department of Biochemistry and Molecular Genetics , U niversity of
Virginia School of Medicine , Charlottesville, V A , U SA
MARJORIE DRAC • Montpellier DNA Combing Facility , B ioCampus Montpellier ,
Montpellier, F rance
KARL E. D UDERSTADT • University of Groningen , G roningen , T he Netherlands
SUSAN L . F ORSBURG • Department of Molecular and Computational Biology , U niversity of
Southern California , L os Angeles, CA , U SA
MARIANA C. GADALETA • Department of Biochemistry and Molecular Biology, D rexel
University College of Medicine , Philadelphia, PA, U SA
HYLKJE J . G EERTSEMA • University of Groningen , G roningen , T he Netherlands
XAVIER G IROUX • Faculté de Médecine Paris Descartes , Université Paris Descartes, Sorbonne
Paris Cité , Paris, France
THIERRY GOSTAN • Institute of Molecular Genetics, CNRS UMR5535 & Université
Montpellier , Montpellier, F rance ; Montpellier DNA Combing Facility, BioCampus
Montpellier , Montpellier, F rance
MARC D . GREEN • Department of Molecular and Computational Biology , University
of Southern California , Los Angeles, C A , USA
ELISABETH GUEYDON • Institute of Molecular Genetics, CNRS UMR5535 & University
Montpellier , Montpellier, F rance
JOYCE L. H AMLIN • Department of Biochemistry and Molecular Genetics, University of
Virginia School of Medicine , C harlottesville, VA, U SA
ix