Novel Strategies to Improve Vaccine Yield Ralph A. Tripp University of Georgia, Athens, GA, USA A major goal is improve influenza (flu) vaccines and vaccine preparedness through the establishment of Vero cell-based enhanced vaccine cell lines, and recombinant flu encoding exogenous miRNA to substantially increase vaccine production. We have identified and validated several host cell genes which upon silencing or knockout (KO) considerably increases flu A and B replication. We use CRISPR-Cas 9 gene editing to generate stable Vero cell KO lines. Vero cell vaccine production has several advantages over traditional egg-based methods, and Vero cells are a FDA-approved cell line used to propagate flu for vaccine production. To further improve flu vaccine production beyond the KO cell lines, we employed reverse genetics technology to express of miRNA in the flu NS1/NS2 splice site to downregulate host genes resulting in faster viral replication and higher HA yield. Together, the flu-miRNA constructs improved flu vaccine production increasing virus titers and HA yield which is central to vaccine dose-sparing and offers lower production costs.
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