This second edition details new and updated methods on circular RNA. Chapter guide readers through circular RNA purification, characterization, circRNA detection, sequence validation, quantification , techniques related to gain- and loss-of-function approaches, circular RNA synthesis, split ligation, engineering, nanoparticle packaging, RNA modifications on circular RNA biogenesis, RNA translation potential, and vaccines based on circular RNAs. Written in the highly successful series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory prools, and key tips on troubleshooting and avoiding known pitfalls.