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DNA Topoisomerase Protocols

  •  ISBN-10 0-89603-444-5

  •  ISBN-13 978-0-89603-444-0

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publication info   M.Bjornsti and N. Osheroff, ed., Humana Press, 1999, 344 pp., hard cover
mfr. no.   Humana

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General description

This is a collection of protocols for investigating DNA structure, topology, and DNA topoisomerase function. These methods utilize new approaches to study changes in DNA topology (linking number, knotting, catenation, and relaxation) and DNA structure (bending), as well as to assess chromosome structure through FLP-mediated recombination and to analyze bacterial nucleoid structure. Protocols detail the expression and purification of bacterial, viral, and eukaryotic DNA topoisomerases.

Table of Contents

Introduction to DNA Topoisomerases. Resolution of DNA Molecules by One-Dimensional Agarose-Gel Electrophoresis,Mary-Ann Bjornsti and Maureen D. Megonigal. Two-Dimensional Agarose-Gel Electrophoresis of DNA Topoisomers, Analysis of Altered DNA Structures: Cruciform DNA, Albert J. Courey. Purification of Supercoiled Plasmid DNA. Purification and Use of DNA Minicircles with Different Linking Numbers. Isolation of Kinetoplast DNA, Isolation of Knotted DNA from Coliphage P4, Analysis of DNA Knots and Catenanes by Agarose-Gel Electrophoresis. Sedimentation Analysis of Bacterial Nucleoid Structure, Coating DNA with RecA Protein to Distinguish DNA Path by Electron Microscopy. Methods for Analyzing DNA Bending. Formation of Extrachromosomal DNA Rings in Saccharomyces cerevisiae Using Site-Specific Recombination. Overexpression and Purification of Bacterial DNA Gyrase, Overexpression and Purification of Bacterial DNA Topoisomerase I. Overexpression and Purification of Escherichia coli DNA Topoisomerase III, Overexpression and Purification of Bacterial DNA Topoisomerase IV, s. Purification of Bacteriophage T4 Type II DNA Topoisomerase, Overexpression and Purification of DNA Topoisomerase I from Yeast, Overexpression and Purification of Saccharomyces cerevisiae DNA Topoisomerase II from Yeast. Purification of DNA Topoisomerase II from Drosophilamelanogaster, S. Purification of DNA Topoisomerase I from Human Placenta. Purification of Baculovirus-Expressed Human DNA Topoisomerase I. Rapid Purification of DNA Topoisomerase II Containing a Hexahistidine Tag by Metal Ion Affinity Chromatography, Metabolic Labeling, Immunoprecipitation, and Two-Dimensional Tryptic Phosphopeptide Mapping of Human Topoisomerase II, Immunoblot Analysis and Band Depletion Assays. Visualization of DNA Topoisomerases by Electron Microscopy, Appendix: Compendium of DNA Topoisomerase Sequences.

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