Teng Michael N, Tran Kim C
Division of Allergy and Immunology, Department of Internal Medicine, University of South Florida Morsani College of Medicine, Tampa, FL, USA.
Methods Mol Biol. 2025;2948:97-107. doi: 10.1007/978-1-0716-4666-3_7.
Minigenome assays have been essential tools in the understanding of viral transcription and RNA replication for respiratory syncytial virus (RSV). Here, we describe the RSV minigenome assay for determining transcription by the viral polymerase in the absence of infection. We detail three different methods of detecting viral RNA synthesis: a firefly luciferase assay for rapid and sensitive measurement of RSV polymerase activity; a dual luciferase assay for assessing contributions of cis-acting signals for RSV transcription; and a real-time quantitative PCR method for determination of specific effects on the transcription of individual viral genes and the polar transcription gradient of RSV.
微型基因组分析一直是理解呼吸道合胞病毒（RSV）病毒转录和RNA复制的重要工具。在此，我们描述了用于在无感染情况下确定病毒聚合酶转录的RSV微型基因组分析。我们详细介绍了三种检测病毒RNA合成的不同方法：一种用于快速灵敏测量RSV聚合酶活性的萤火虫荧光素酶分析；一种用于评估顺式作用信号对RSV转录贡献的双荧光素酶分析；以及一种用于确定对单个病毒基因转录的特定影响和RSV极性转录梯度的实时定量PCR方法。
