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Updated: Jan 4, 2026
Simplified Reverse Genetics Method to Recover Recombinant Rotaviruses Expressing Reporter Proteins
Published on: April 17, 2020
Gene-edited vero cells as rotavirus vaccine substrates
Nichole Orr-Burks1, Jackelyn Murray1, Weilin Wu1
1Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA.
Background:
Rotavirus (RV) is a leading cause of severe gastroenteritis globally and can cause substantial morbidity associated with gastroenteritis in children <5 years of age. Orally administered live-attenuated RV vaccines offer protection against disease but vaccination efforts have been hampered by high manufacturing costs and the need to maintain a cold chain.
Methods:
A subset of Vero cell host genes was identified by siRNA that when knocked down increased RV replication and these anti-viral host genes were individually deleted using CRISPR-Cas9.
Results:
Fully-sequenced gene knockout Vero cell substrates were assessed for increased RV replication and RV vaccine antigen expression compared to wild type Vero cells. The results showed that RV replication and antigen production were logs higher in Vero cells having an EMX2 gene deletion compared to other Vero cell substrates tested.
Conclusions:
We used siRNAs to screen for host genes that negatively affected RV replication, then CRISPR-Cas9 gene editing to delete select genes. The gene editing led to the development of enhanced RV vaccine substrates supporting a potential path forward for improving RV vaccine production.
