This detailed new edition collects cutting-edge laboratory protocols, techniques, and applications in use by some of the leading international experts in the broad field of chromosome architecture. The book emphasizes the increasing physiological relevance of chromosome architecture investigation, manifest both through application of more complex bottom-up assays in vitro as well as through maintaining the native physiological context through the investigation of living, functional cells. In addition, the chapters reflect the dramatic improvement in the length scale of precision by utilizing single-molecule approaches, both for imaging the DNA content of chromosome and proteins that bind to DNA as well as using methods that can controllably manipulate single DNA molecules, and the use of advanced computational methods and mathematical analysis is also featured. Written for 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, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and up-to-date, Chromosome Architecture: Methods and Protocols, Second Edition is an ideal guide for researchers working in this dynamic area of study.

A Next Generation of Advances in Chromosome Architecture.- Single Molecule Narrowfield Microscopy of Protein-DNA Binding Dynamics in Glucose Signal Transduction of Live Yeast Cells.- Convolutional Neural Networks for Classifying Chromatin Morphology in Live Cell Imaging.- Fluorescence Recovery After Photobleaching (FRAP) to Study Dynamics of the Structural Maintenance of Chromosome (SMC) Complex in Live Escherichia coli Bacteria.- Atomic Force Microscopy of DNA and DNA-Protein Interactions.- Live-Cell Visualization of DNA Transfer and Pilus Dynamics during Bacterial Conjugation.- Transverse Magnetic Tweezers Allowing Coincident Epi-Fluorescence Microscopy on Horizontally Extended DNA.- Atomistic Molecular Dynamics Simulations of DNA in Complex 3D Arrangements for Comparison with Lower Resolution Structural Experiments.- Replication Labeling Methods for Super-Resolution Imaging of Chromosome Territories and Chromatin Domains.- DNA-Protein Interactions Studied Directly Using iSCATImaging of GNP-Tagged Proteins.- Escherichia coli Chromosome Copy Number Measurement Using Flow Cytometry Analysis.- Dynamics of Bacterial Chromosomes by Locus Tracking in Fluorescence Microscopy.- Measuring Nuclear Mechanics with Atomic Force Microscopy.- Copy Number Analysis of the Yeast Histone Deacetylase Complex Component Cti6 Directly in Living Cells.- Super-Resolution Microscopy and Tracking of DNA-Binding Proteins in Bacterial Cells.- Studying the Dynamics of Chromatin-Binding Proteins in Mammalian Cells Using Single-Molecule Localization Microscopy.- The End Restraint Method for Mechanically Perturbing Nucleic Acids In Silico.- Visualizing the Replisome, Chromosome Breaks, and Replication Restart in Bacillus subtilis.- High-Throughput Imaging of Bacillus subtilis.- Measuring Nuclear Organization of Proteins with STORM Imaging and Cluster Analysis.- Single-Molecular Quantification of Flowering Control Proteins within Nuclear Condensates in Live Whole Arabidopsis Root.