Department of Population Health, Poultry Diagnostic and Research Center, University of Georgia, Athens, GA, USA.
Food Animal Health Research Program, Department of Veterinary Preventive Medicine, Ohio Agricultural Research and Development Center, College of Veterinary Medicine, The Ohio State University, Wooster, OH, USA.
Methods Mol Biol. 2020;2123:37-59. doi: 10.1007/978-1-0716-0346-8_4.
Reverse genetics is the process of generating an RNA virus from a cDNA copy. Reverse genetics systems have truly transformed our ability to manipulate and study negative-strand RNA viruses. Plasmid-based reverse genetics approaches for influenza viruses provide a better understanding of virulence, transmission, mechanisms of antiviral resistance, and the development of alternative vaccines and vaccination strategies. Studying the molecular changes that allow influenza A viruses (IAVs) to transmit among animal species is important to better understand their animal health and public health risks. In this chapter, the cloning of cDNA copies of IAV's RNA segments into a reverse genetics plasmid vector, the experimental procedures for studying viral polymerase activity, and the successful generation of recombinant IAVs are described.
反向遗传学是从 cDNA 拷贝生成 RNA 病毒的过程。反向遗传学系统真正改变了我们操纵和研究负链 RNA 病毒的能力。流感病毒的基于质粒的反向遗传学方法提供了对毒力、传播、抗病毒耐药机制以及替代疫苗和接种策略的发展的更好理解。研究允许甲型流感病毒 (IAV) 在动物物种之间传播的分子变化对于更好地了解它们对动物健康和公共卫生的风险非常重要。在本章中，描述了将 IAV 的 RNA 片段的 cDNA 拷贝克隆到反向遗传学质粒载体中、研究病毒聚合酶活性的实验程序以及成功生成重组 IAV 的过程。
