Construction of BHV-1 UL41 Defective Virus Using the CRISPR/Cas9 System and Analysis of Viral Replication Properties

Construction of BHV-1 UL41 Defective Virus Using the CRISPR/Cas9 System and Analysis of Viral Replication Properties

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Bovine herpesvirus type 1 (BHV-1) is a neurotropic herpesvirus that causes infectious rhinotracheitis and vulvovaginitis in cattle.The virion host shutoff protein encoded by the BHV-1 UL41 gene is highly conserved in the Alphaherpesvirinae subfamily.This protein can degrade viral and host messenger RNA (mRNA) to interrupt host defense and Stretch Mark Butter facilitate the rapid proliferation of BHV-1.

However, studies on the BHV-1 UL41 gene are limited, and BHV-1 defective virus construction using the CRISPR/Cas9 system is somewhat challenging.In this study, we rapidly constructed a BHV-1 UL41-deficient strain using the CRISPR/Cas9 system in BL primary bovine-derived cells.BHV-1 UL41-defective mutants were screened by Western blot analysis using specific polyclonal antibodies as the primary antibodies.

During the isolation and purification of the defective strain, a mixed virus pool edited by an efficient single-guide RNA (sgRNA) showed a plaque number reduction.Viral growth property assessment showed that BHV-1 UL41 was dispensable for replication, but the UL41-defective strain exhibited early and slowed viral replication.Furthermore, the BHV-1 UL41-deficient strain exhibited ONION CUBES enhanced sensitivity to temperature and acidic environments.

The BHV-1 UL41-deficient strain regulated viral and host mRNA levels to affect viral replication.