Introduction 384 2. Coronavirus reverse genetics systems 386 2.1 Reverse genetic system using targeted RNA recombination 387 2.2 Reverse genetic system using in vitro ligation 387 2.3 Reverse genetic system using bacteria artificial chromosomes 390 2.4 Reverse genetic system using transformation associated recombination cloning 390 2.5 Reverse genetic system using vaccinia virus vectors 392 2.6 Rapid gene manipulation of coronavirus 392 3. Rational design of vaccine candidates using reverse genetics technology 393 3.1 Live attenuated vaccines 393 3.2 Recombinant vector vaccines 403 4. Study the cell and tissue tropism of coronaviruses using reverse genetics 404 5. Screening antiviral drugs using reverse genetics 405 6. Conclusion and future directions 405 Acknowledgments 406 References 406

CoVs can spill from an animal reservoir into a naive host to cause diseases in humans or domestic animals. PEDV results in high mortality in piglets, which has caused immense economic losses in the pork industry.