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BKV infection or reactivation may cause BKV\u2010associated nephropathy and hemorrhagic cystitis in immunosuppressed transplant recipients. No effective antivirals or prevention strategies are available against BKV infections. The current BKV reverse system employs the transfection of purified full\u2010length linear viral genomes released by enzyme digestion from BKV genomic plasmids. The method is laborious and often results in variable DNA yield and quality, which can affect the efficiency of transfection and subsequent formation of circular viral genomes in cells. In this study, we report the generation of circular viral genomes by Cre\u2010mediated DNA recombination in cells directly transfected with BKV precursor genomic plasmids. The novel system supported efficient viral expression and replication, and produced a higher level of infectious virions compared with the transfection with linear BKV genomes. Furthermore, we successfully constructed recombinant BKV capable of reporter gene expression. In conclusion, the novel BKV reverse genetic system allows for simpler manipulation of BKV genome with better virus yield, providing a tool for the study of BKV life cycle and antiviral screening.<\/jats:p>","DOI":"10.1002\/jmv.28995","type":"journal-article","created":{"date-parts":[[2023,7,31]],"date-time":"2023-07-31T08:50:51Z","timestamp":1690793451000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Establishment and characterization of a novel reverse genetic system of BK polyomavirus"],"prefix":"10.1002","volume":"95","author":[{"given":"Nannan","family":"Liu","sequence":"first","affiliation":[{"name":"Key Laboratory of Medical Molecular Virology (MOE\/NHC\/CAMS), Shanghai Institute of Infectious Diseases and Biosecurity, School of Basic Medical Sciences, Shanghai Medical College Fudan University Shanghai 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