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Microbiol."],"abstract":"<jats:p>The mini-genome reporter assay is a key tool for conducting RNA virus research. However, procedural complications and the lack of adequate literature pose a major challenge in developing these assay systems. Here, we present a novel, yet generic and simple, cloning strategy for the construction of an influenza B virus reporter RNA template and describe an extensive standardization of the reporter RNP\/polymerase activity assay for monitoring viral RNA synthesis in an infection-free setting. Using this assay system, we showed for the first time the effect of viral protein NS1 and host protein kinase C delta (PKCD) on influenza B virus RNA synthesis. In addition, the assay system showed promising results in evaluating the efficacy of antiviral drugs targeting viral RNA synthesis and virus propagation. Together, this work offers a detailed protocol for the standardization of the influenza virus minigenome assay and an excellent tool for screening of host factors and antivirals in a fast, user-friendly, and high-throughput manner.<\/jats:p>","DOI":"10.3389\/fmicb.2022.868367","type":"journal-article","created":{"date-parts":[[2022,5,25]],"date-time":"2022-05-25T09:33:39Z","timestamp":1653471219000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":2,"title":["A Comprehensive Roadmap Towards the Generation of an Influenza B Reporter Assay Using a Single DNA Polymerase-Based Cloning of the Reporter RNA Construct"],"prefix":"10.3389","volume":"13","author":[{"given":"Nandita","family":"Kedia","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Saptarshi","family":"Banerjee","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Arindam","family":"Mondal","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2022,5,25]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","first-page":"81","DOI":"10.4161\/hv.8.1.17623","article-title":"The rationale for quadrivalent influenza vaccines","volume":"8","author":"Ambrose","year":"2012","journal-title":"Hum. 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