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

Blog Article

Bovine herpesvirus type 1 (BHV-1) is a neurotropic herpesvirus that causes infectious rhinotracheitis and vulvovaginitis in cattle.The virion host shutoff protein telemarkskongen flue 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 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 3m speedglas 9002nc 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 enhanced sensitivity to temperature and acidic environments.The BHV-1 UL41-deficient strain regulated viral and host mRNA levels to affect viral replication.