DNA Topoisomerases

by Clarke, Duncan J.

• ISBN13: 9781607613398
• ISBN10: 1607613395
• Edition: 1st
• Format: Hardcover
• Copyright: 2009-12-04
• Publisher: Humana Pr Inc

Summary

Due to their importance to cell biology, DNA topology-altering enzymes, collectively DNA topoisomerases and gyrases, have been studied thoroughly from a biochemical perspective, yet the expansion of research at the molecular and cellular levels continues to show a great deal of promise. In DNA Topoisomerases: Methods and Protocols, experts in the field present a compendium of protocols that extends into the new areas of research that have been recently uncovered in order to elucidate the methodology needed to study topoisomerases in the molecular and cellular context. The volume examines subjects such as the analysis of DNA topologies and the interaction of topoisomerases with DNA as tools to measure topoisomerase functions, the binding of topoisomerases to specific sites in the genomes of eukaryotes, including origins of DNA replication, as well as exploration of perturbed topoisomerase function. As a volume in the highly successful Methods in Molecular Biology'_¢ series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, laboratory protocols, and notes on trouble-shooting and avoiding known pitfalls.Cutting-edge and easy-to-use, DNA Topoisomerases: Methods and Protocols provides key content which reflects an expanded understanding of the functions of these essential enzymes in the hope of inspiring researchers to further important discoveries in this dynamic field.

Table of Contents

Preface	p. v
Contributors	p. ix
Introduction: Emerging Themes in DNA Topoisomerases Research	p. 1
Analysis of DNA Topoisomers, Knots, and Catenanes by Agarose Gel Electrophoresis	p. 11
Two-Dimensional Agarose Gel Electrophoresis of DNA Topoisomers	p. 27
Cleavage of Plasmid DNA by Eukaryotic Topoisomerase II	p. 39
Assays for the Preferential Binding of Human Topoisomerase I to Supercoiled DNA	p. 49
Monitoring the Topoisomerase II DNA Gate Conformational Change with Fluorescence Resonance Energy Transfer	p. 59
Single-Molecule Magnetic Tweezers Studies of Type IB Topoisomerases	p. 71
Dissolution of Double Holliday Junctions by the Concerted Action of BLM and Topoisomerase II¿	p. 91
ChIP-on-Chip Analysis of DNA Topoisomerases	p. 103
Analyzing Top2 Distribution on Yeast Chromosomes by Chromatin Immunoprecipitation	p. 119
Binding of DNA Topoisomerases I and II to Replication Origins	p. 131
Cytometric Assessment of DNA Damage Induced by DNA Topoisomerase Inhibitors	p. 145
Analysis of the Topoisomerase II-Dependent Decatenation G2 Checkpoint and Checkpoint Kinases in Human Cells	p. 155
Assaying Topoisomerase II Checkpoints in Yeast	p. 167
Cytological Analysis of Chromosome Structural Defects that Result from Topoisomerase II Dysfunction	p. 189
Top2 SUMO Conjugation in Yeast Cell Lysates	p. 209
Analysis of SUMOylation of Topoisomerase II¿ with Xenopus Egg Extracts	p. 221
The Dynamics of DNA Topoisomerase II¿ in Living Cells	p. 233
Deplection and Mutation of Topoisomerase II in Animal Cells	p. 245
Subject Index	p. 265
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