Ray H. Gavin brings together an international panel of experienced researchers to detail the readily reproducible methods that utilize biochemistry, immunology, genetics, microscopy, and image analysis for investigating cytoskeleton structure and function. Each protocol contains proven step-by-step instructions that enable both the novice and the experienced researcher to achieve successful experimental results. The protocols utilize diverse model systems in a variety of organisms, including Saccharomyces, Micrasterias, Tetrahymena, Drosophila, Spisula, and Xenopus. Microscopy applications include digital-video microscopy and computer-assisted systems for the evaluation of cell motility and morphology. State-of-the-art and highly practical, Cytoskeleton Methods and Protocols makes available a diverse collection of powerful experimental systems and tools for successfully studying cytoskeleton structure and function.
Part I. Identification of Cytoskeleton Proteins. Using an Inverse PCR Strategy to Clone Large, Contiguous Genomic DNA Fragments, Jorge A. Garces and Ray H. Gavin. Microsequencing of Myosins for PCR Primer Design, Elaine L. Bearer. Evaluating the Dynein Heavy Chain Gene Family in Tetrahymena,Gangadhara Sailaja, Leslie M. Lincoln, Jifan Chen, and David J. Asai. Part II. Microscopy Applications. Video-Enhanced Microscopy for Analysis of Cytoskeleton Structure and Function, George M. Langford. Computer-Assisted Systems for the Analysis of Amoeboid Cell Motility, David R. Soll, Deborah Wessels, Edward Voss, and Olof Johnson. Evaluation of Individual-Cell Motility, Peter S. Walmod, Rasmus Hartmann-Petersen, Anton Berezin, Soren Prag, Vladislav V. Kiselyov, Vladimir Berezin, and Elizabeth Bock. Evaluation of Cell Morphology, Eugene A. Lepekhin, Peter S. Walmod, Anton Berezin, Vladimir Berezin, and Elisabeth Bock. A Quantitative Assay for Measurement of Chemokinesis in Tetrahymena, Uffe Koppelhus, Per Hellung-Larsen, and Vagn Leick. Part III. Reagents for Studying Cytoskeleton Protein Function. Jasplakinolide: An Actin-Specific Reagent that Promotes Actin Polymerization, Andreas Holzinger. Site-Directed Antibodies as Tools For Investigating Structure and Function of Cytoskeleton Proteins, Walter Steffen and Julie L. Hodgkinson. Direct Labeling of Components in Protein Complexes by Immuno-Electron Microscopy, Julie L. Hodgkinson and Walter Steffen. Tat-Mediated Delivery of Antibodies into Cultured Cells, Walter Steffen. Part IV. Cytoskeleton Dynamics. Studying Cytoskeletal Dynamics in Living Cells Using Green Fluorescent Protein, Yisang Yoon, Kelly R. Pitts, and Mark A. McNiven. Use of Green Fluorescent Protein to Study Cellular Dynamics: Constructing GFP-Tagged Motor Enzymes, Hong Cao, Heather M. Thompson, Eugene W. Krueger, and Mark A. McNiven. Transient Transfections and Heterokaryons as Tools for the Analysis of Keratin IF Dynamics, Jesus M. Paramio and Jose L. Jorcano. Part V. Cellular Systems as Tools for Investigating Cytoskeleton Structure and Function. Chromatophores as Tools for the Study of Organelle Transport, Bruce R. Telzer and Leah T. Haimo. Centriole Duplication, Centrosome Maturation, and Spindle Assembly in Lysates of Spisula solidissima Oocytes, Andrew W. Suddith, Eugeni A. Vaisberg, Sergei A. Kuznetsov, Walter Steffen, Conly L. Rieder, and Robert E. Palazzo. Xenopus Egg Extracts as a Model System for Analysis of Microtubule, Actin Filament, and Intermediate Filament Interactions, Craig A. Mandato, Kari L. Weber, Anna J. Zandy, Thomas J. Keating, and William M. Bement. Detergent-Extracted Models for the Study of Cilia or Flagella, Charles B. Lindemann and Kathleen A. Schmitz. Part VI. Genetic Approaches for Studies of Cytoskeleton Protein Function. A Yeast Two-Hybrid Approach for Probing Cytoskeletal Protein Interactions, Jin-jun Meng and Wallace Ip. Manipulating Dynein Genes in Tetrahymena, David J. Asai, K. Mark DeWall, Leslie M. Lincoln, and Renotta K. Smith. Functional Analysis of Cytoskeletal Components in the Developing Visual System of Drosophila, Qi He. Index.