Dictyostelium discoideum Protocols

Dictyostelium discoideum is a well-establish eukaryotic model organism that offers unique advantages for studying fundamental cellular processes, including signal transduction, random and directed cell motility, cytokinesis, endocytosis and vesicle transport and development. Dictyostelium is also increasingly used for the investigation of human disease genes and the crosstalk between host and pathogen. The availability of the genome sequence at a mouse click together with a whole range of supporting information and resources along with a powerful armoury of molecular genetics techniques have considerably enhanced the experimental attractiveness of D. discoideum in recent years.

The second edition of Dictyostelium discoideum Protocols incorporates the most recent developments in a number of fields. The book is divided into four parts. The first part provides an introduction to the amoebozoa and community resources. The second part presents large-scale analysis methods made possible by the completion of the Dictyostelium genome sequence. The third part is dedicated to molecular genetics techniques, cell biological, biochemical and biophysical methods. The chapters in the fourth part describe the use of Dictyostelium as a model system to study vesicle formation, trafficking and infection by bacterial pathogens.

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 key tips on troubleshooting and avoiding known pitfalls.

 Authoritative and practical, Dictyostelium discoideum Protocols, Second Edition seeks to aid scientists in the application of innovating techniques to study a range of fundamental biological processes in this attractive model organism.

1. The Amoebozoa

Christina Schilde and Pauline Schaap

Salvatore Bozzaro

4. One Stop Shop for Everything Dictyostelium: DictyBase and the Dicty Stock Center in 2012

Petra Fey, Robert J. Dodson, Siddhartha Basu and Rex L. Chisholm

5. Fluorescent Reporters and Methods to Analyze Fluorescent Signals

6. Collection and Cultivation of Dictyostelids from the Wild

Tracy E. Douglas, Debra A. Brock, Boahemaa Adu-Oppong, David C. Queller and Joan E. Strassmann

7. Identification and Verification of microRNAs by High-throughput Sequencing

Jimmie Hällman, Lotta Avesson, Johan Reimegård, Max Käller and Fredrik Söderbom

8. Transcriptional Profiling of Dictyostelium with RNA Sequencing

Edward Roshan Miranda, Gregor Rot, Marko Toplak, Balaji Santhanam, Tomaz Curk, Gad Shaulskyand Blaz Zupan

9. Analysis of Chromatin Organization by Deep Sequencing Technologies

James L. Platt, Nick A. Kent, Adrian J. Harwood, and Alan R. Kimmel

10. Pharmacogenetics of Resistance to Cisplatin and Other Anti-cancer Drugs and the Role of Sphingolipid Metabolism

Stephen Alexander, William S. Swatson, Hannah Alexander

11. N-glycomic and –Glycoproteomic Studies in the Social Amoebae

Christa L. Feasley, Alba Hykollari, Katharina Paschinger, Iain B. H. Wilson, and Christopher M. West

12. Measuring Cheating, Fitness, and Segregation in D. discoideum

Neil J Buttery, Jeff smith, David C Queller and Joan E Strassmann

13. The Application of the Cre-loxP System for Generating Multiple Knock-Out and Knock-In Targeted Loci

Jan Faix, Joern Linkner, Benjamin Nordholz, James L. Platt, Xin-Hua Liao, and Alan R. Kimmel

Douwe M. Veltman¹ and Peter J.M. Van Haastert²

15. Isolation of Dictyostelium Nuclei for Light and Electron Microscopy

Petros Batsios, Otto Baumann, Ralph Gräf and Irene Meyer

16. Investigation of DNA Repair Pathway Activity

Anne-Marie C. Couto, Nicholas D. Lakin and Catherine J. Pears

17. Transcript Localization in D. discoideum Cells by RNA FISH

Patrick Hofmann, Janis Kruse and Christian Hammann

Jessica E. Accari, Sam Manna, Paul R. Fisher, and Christian Barth

Sarah J. Annesley, Sergio Carilla-Latorre, Ricardo Escalante and Paul R. Fisher

20. Micropipette Aspiration for Studying Cellular Mechanosensory Responses and Mechanics

Yee-Seir Kee and Douglas N. Robinson

21. Quantitative Analysis of Phagocytosis and Phagosome Maturation

Natascha Sattler, Roger Monroy and Thierry Soldati

22. Setting Up and Monitoring an Infection of Dictyostelium discoideum with Mycobacteria

Sonia Arafah, Sébastien Kicka, Valentin Trofimov, Monica Hagedorn, Nuria Andreu, Siouxsie Wiles, Brian Robertson, and Thierry Soldati

23. Isolation of Pathogen-containing Vacuoles

Olga Shevchuk and Michael Steinert

24. Immuno-magnetic Purification of Fluorescent Legionella-containing Vacuoles

Ivo Finsel, Christine Hoffmann and Hubert Hilbi

25. Secretory Lysosomes in Dictyostelium: Visualization, Characterization and Dynamics

Wanessa C. Lima and Pierre Cosson

26. Monitoring Autophagy in Dictyostelium

Ana Mesquita, Javier Calvo-Garrido, Sergio Carilla-Latorre and Ricardo Escalante