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Published on: January 7, 2019
IRF7-deficient MDBK cell based on CRISPR/Cas9 technology for enhancing IBRV replication
Guiyang Ge1, Dongli Li2, Qian Ling3
1College of Animal Science and Technology, Jilin Agricultural University, Changchun, China.
Insights
Researchers developed IRF7-deleted MDBK cells to improve infectious bovine rhinotracheitis (IBR) vaccine production. These modified cells enhance bovine alphaherpesvirus-1 (BoAHV-1) replication, enabling the creation of higher-titer vaccines.
Area of Science:
- Veterinary Virology
- Cell Biology
- Immunology
Background:
- Infectious bovine rhinotracheitis (IBR), caused by bovine alphaherpesvirus-1 (BoAHV-1), is a significant cattle disease managed by vaccination.
- Madin-Darby bovine kidney (MDBK) cells are crucial for IBR vaccine production but are subject to innate immune gene inhibition of viral replication.
Purpose of the Study:
- To develop IRF7-deleted MDBK cells to enhance bovine alphaherpesvirus-1 (BoAHV-1) replication.
- To facilitate the production of high-titer infectious bovine rhinotracheitis (IBR) vaccines.
Main Methods:
- CRISPR/Cas9 technology was employed to knockout the IRF7 gene in MDBK cells.
- Virus growth curves, CCK-8 assays, cell scratch assays, and qPCR were used to assess viral replication and gene expression.
Main Results:
- IRF7 knockout in MDBK cells significantly increased IBRV replication capacity.
- A notable reduction in type I interferons (IFN-α and IFN-β) expression was observed in IRF7 knockout cells.
- The IRF7-/- MDBK cell lines demonstrated enhanced production of high-titer IBRV.
Conclusions:
- IRF7 knockout MDBK cell lines are effective for high-titer IBRV production.
- This advancement supports the development of more effective inactivated or attenuated IBR vaccines.
Abstract:
Infectious bovine rhinotracheitis (IBR), characterized by acute respiratory lesions in cattle, is a major infectious disease caused by bovine alphaherpesvirus-1 (BoAHV-1). Control of this disease is primarily depending on vaccination. Madin-Darby bovine kidney cells (MDBK) being the main host cells and the important production platform for IBR vaccines. However, innate immune genes inhibit viral replication. Accordingly, the aim of this study was developing of IRF7 gene deleted MDBK cells to facilitate the production of high-titer vaccines. The CRISPR/Cas9 technology was used to knock out the IRF7 gene in MDBK cells and the impact on virus replication was examined using virus growth curves, CCK-8 assays, cell scratch assays, and qPCR. The knockout of the IRF7 gene in MDBK cells led to an increased replication capacity of IBRV and a significant reduction in type I interferons expression, specifically IFN-α and IFN-β. This indicates that IRF7 -/-MDBK cell lines can effectively result in production of IBRV with high-titer, which will enhance the development of inactivated or attenuated vaccines.
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