This volume aims to describe a variety of techniques that reflects the wide range of research currently performed in the field of coronavirology, and begins with an overview of current understandings of coronavirus replication and pathogenesis to introduce specialists and non-specialists to the field. The rest of the book is divided into several sections of chapters beginning with those that describe identification, diagnosis, and study of the evolution of coronaviruses. The next few chapters discuss the preparation of cells and organ cultures useful in propagating coronaviruses and titration techniques, as well as techniques for analyzing virus functions that require purification of the viruses. The next chapters describe two commonly used reverse genetics techniques for coronaviruses, and techniques detailing identification of cellular receptors, binding profiles of viral attachment proteins, and virus-cell fusion. The final chapters cover a broad spectrum of techniques to identify virus-host protein-protein interactions, confirm the functional role of these proteins in virus replication, study host cell responses through genome-wide or pathway-specific approaches, and visualize virus replication complexes. Written in the highly successful Methods in Molecular Biology series format, the chapters include the kind of detailed description and implementation advice that is crucial for getting optimal results in the laboratory.Authoritative and practical, Coronaviruses: Methods and Protocols appeals to a wide variety of scientists because it highlights techniques that are currently used in the coronovirology field, while also discussing practices applicable to other virology fields.

Includes cutting-edge methods and protocols

1. Coronaviruses: An Overview of Their Replication and PathogenesisAnthony R. Fehr and Stanley PerlmanSection I: Detection, Diagnosis and Evolution of Coronaviruses2. Identification of a Novel Coronavirus from Guinea Fowl Using MetagenomicsMariette F. Ducatez and Jean-Luc Guérin3. Serological Diagnosis of Feline Coronavirus Infection by Immunochromatographic TestTomomi Takano and Tsutomu Hohdatsu4. Estimation of Evolutionary Dynamics and Selection Pressure in CoronavirusesMuhammad Munir and Martí CorteySection II: Propogation, Titration, and Purification of Coronaviruses5. The Preparation of Chicken Tracheal Organ Cultures for Virus Isolation, Propagation, and TitrationRuth M. Hennion6. The Preparation of Chicken Kidney Cell Cultures for Virus PropagationRuth M. Hennion and Gillian Hill7. Isolation and Propagation of Coronaviruses in Embryonated EggsJames S. Guy8. Characterization of Human Coronaviruses on Well-Differentiated Human Airway Epithelial Cell CulturesHulda R. Jonsdottir and Ronald Dijkman9. Quantification of Infectious Bronchitis Coronaviruses by Titration in vitro and in ovoJoeri Kint, Helena J. Maier, and Erik Jagt10. Purification of Coronavirus Virions for Cryo-EM and Proteomic AnalysisStuart Dent and Benjamin W. Neuman11. Partial Purification of IBV and Subsequent Isolation of Viral RNA for Next Generation SequencingSarah M. Keep, Erica Bickerton, and Paul BrittonSection III: Manipulating the Genomes of Coronaviruses12. Transient Dominant Selection for the Modification and Generation of Recombinant Infectious Bronchitis CoronavirusesSarah M. Keep, Erica Bickerton, and Paul Britton13. Engineering Infectious cDNAs of Coronavirus as Bacterial Artificial ChromosomesFernando Almazán, Silvia Márquez-Jurado, Aitor Nogales, and Luis EnjuanesSection IV: Coronavirus Attachment and Entry14. Protein Histochemistry Using Coronaviral Spike Proteins: Studying Binding Profiles and Sialic Acid Requirements for Attachment to TissuesI. N. Ambepitiya Wickramasinghe, M. H. Verheije15. Identification of Protein Receptors for Coronaviruses by Mass Spectrometry V. Stalin Raj, Mart M. Lamers, Saskia L. Smits, Jeroen A. A. Demmers, Huihui Mou, Berend-Jan Bosch, Bart L. Haagmans16. Single Particle Tracking Assay to Study Coronavirus Membrane FusionDeirdre A. Costello and Susan DanielSection V: Studying Virus-Host Interactions17. Studying Coronavirus-Host Protein InteractionsChee-Hing Yang, Hui-Chun Li, Cheng-Huei Hung, and Shih-Yen Lo18. A Field-Proven Yeast Two-Hybrid Protocol Used to Identify Coronavirus-Host Protein-Protein InteractionsPierre-Olivier Vidalain, Yves Jacob, Marne C. Hagemeijer, Louis M. Jones, Grégory Neveu, Jean-Pierre Roussarie, Peter J. M. Rottier, Frédéric Tangy, and Cornelis A. M. de Haan19. Investigation of the Functional Roles of Host Cell Proteins Involved in Coronavirus Infection Using Highly Specific and Scalable RNA Interference (RNAi) ApproachJean Kaoru Millet and Béatrice Nal20. Transcriptome Analysis of Feline Infectious Peritonitis Virus (FIPV) InfectionParvaneh Mehrbod, Mohammad Syamsul Reza Harun, Ahmad Naqib Shuid, Abdul Rahman Omar21. Quantification of Interferon Signaling in Avian CellsJoeri Kint and Maria Forlenza22. Studying the Dynamics of Coronavirus Replicative StructuresMarne C. Hagemeijer and Cornelis A. M. de Haan23. Preparation of Cultured Cells Using High Pressure Freezing and Freeze Substitution for Subsequent 2D or 3D Visualization in the Transmission Electron MicroscopePhilippa C. Hawes