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Stable expression of shRNA for the control of recombinant adenovirus replication

Preventing the replication of adenovirus could have practical uses, such as controlling infection with wild-type virus or in applications involving recombinant vectors. Mainly transient methods have been used to inhibit adenovirus replication, including siRNA or drugs. Here, we tested whether stable...

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Detalles Bibliográficos
Autores principales: Lana, M.V.G., Antunes, F., Tessarollo, N.G., Strauss, B.E.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Associação Brasileira de Divulgação Científica 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10361640/
https://www.ncbi.nlm.nih.gov/pubmed/37493770
http://dx.doi.org/10.1590/1414-431X2023e12682
Descripción
Sumario:Preventing the replication of adenovirus could have practical uses, such as controlling infection with wild-type virus or in applications involving recombinant vectors. Mainly transient methods have been used to inhibit adenovirus replication, including siRNA or drugs. Here, we tested whether stable expression of shRNA designed to target hexon, Iva2, or pol can inhibit the replication of a recombinant adenoviral vector, Ad-LacZ (serotype 5, E1/E3 deleted), in 293T cells. Significant knockdown correlating with reduced Ad-LacZ replication was achieved only when hexon was targeted. Cell sorting and isolation of cellular clones further accentuated knockdown of the hexon transcript, reduced protein levels by more than 90%, and diminished adenovirus production. As visualized by transmission electron microscopy, the cellular clone expressing the hexon-specific shRNA yielded 89.2% fewer particles compared to the parental 293T cells. Full scale production followed by purification revealed a 90.2% reduction in Ad-LacZ biological titer. These results support the notion that stable expression of shRNA can be used as a means to control adenovirus replication.