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This book brings together a unique collection of protocols that cover novel and specialized techniques as well as updated and improved adaptations of more standard procedures involving the cell cycle and its regulation by oscillatory networks, exploring recent progress in the field from both holistic and reductionist perspectives. The edition provides a space for researchers to highlight and explore the latest developments in molecular biology and biochemical techniques for studying oscillatory networks and to share these across the research community to facilitate further progress. Written in the highly successful Methods in Molecular Biology series format, 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, Cell Cycle Oscillators: Methods and Protocols, Second Edition serves as an invaluable guide for those new to the field as well as the more experienced scientist.
Chapters 19 and 21 are available open access under a Creative Commons Attribution 4.0 International License via link.springer.com.
List of contents
Cell Cycle Control: A System of Interlinking Oscillators.- Substrate Phosphorylation Rates as an In Vivo Measurement of Kinase Activity.- Real Time Monitoring of APC/C-Mediated Substrate Degradation Using Xenopus laevis Egg Extracts.- Fluorescent Peptide Biosensors for Probing CDK Kinase Activity in Cell Extracts.- Phosphatase and Kinase Substrate Specificity Profiling with Pooled Synthetic Peptides and Mass Spectrometry.- Whole-Mount Immunostaining for the Identification of Histone Modifications in the S-Phase Nuclei of Arabidopsis Roots.- Construction of a Full-Length 3'UTR Reporter System for Identification of Cell-Cycle Regulating MicroRNAs.- Purification of Cyclin-Dependent Kinase Fusion Complexes for In Vitro Analysis.- Optimizing Cell Synchronization Using Nocodazole or Double Thymidine Block.- Highly Synchronous Mitotic Progression in Schizosaccharomyces pombe Upon Relief of Transient Cdc2-asM17 Inhibition.- Elucidating Human Mitosis Using an Anaphase-Like Cell-Free System.- EDU (5-Ethynyl-2'-Deoxyuridine)-Coupled-Fluorescence-Intensity Analysis: Determining Absolute Parameters of the Cell Cycle.- High-Resolution Analysis of Centrosome Behavior During Mitosis.- Assaying Cell Cycle Progression via Flow Cytometry in CRISPR/Cas9-Treated Cells.- Use of Mitotic Protein Kinase Inhibitors and Phospho-Specific Antibodies to Monitor Protein Phosphorylation During the Cell Cycle.- Visualization of Radiation-Induced Cell Cycle Kinetics with a Fluorescent Ubiquitination-Based Cell Cycle Indicator (Fucci).- Dynamic Behavior of Inactive X Chromosome Territory During the Cell Cycle as Revealed by H3K27me3-Specific Intracellular Antibody.- Analyzing Centrioles and Cilia by Expansion Microscopy.- Analysis of Cell Cycle Progression in the Budding Yeast S. cerevisiae.- Application of PALM Super Resolution Microscopy to the Analysis of Microtubule-Organizing Centers (MTOCs) in Aspergillus nidulans.- Live Imaging and Analysis of Cilia and Cell Cycle Dynamics with the Arl13bCerulean-Fucci2a Biosensor and Fucci Tools.- Calorimetric Heat Dissipation Measurements of Developing Zebrafish Embryos.- The Conditional Knockout Analogous System: CRISPR-Mediated Knock Out Together with Inducible Degron and Transcription-Controlled Expression.
