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CRISPR/Cas9 Mutagenesis of UL21 in Multiple Strains of Herpes Simplex Virus Reveals Differential Requirements for pUL21 in Viral Replication

Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON K7L 3N6, Canada
Author to whom correspondence should be addressed.
Viruses 2018, 10(5), 258;
Received: 26 April 2018 / Revised: 10 May 2018 / Accepted: 13 May 2018 / Published: 15 May 2018
(This article belongs to the Special Issue Applications of CRISPR Technology in Virology 2018)
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Studies from multiple laboratories using different strains or species of herpes simplex virus (HSV) with deletions in UL21 have yielded conflicting results regarding the necessity of pUL21 in HSV infection. To resolve this discrepancy, we utilized CRISPR/Cas9 mutagenesis to isolate pUL21 deficient viruses in multiple HSV backgrounds, and performed a side-by-side comparison of the cell-to-cell spread and replication phenotypes of these viruses. These analyses confirmed previous studies implicating the involvement of pUL21 in cell-to-cell spread of HSV. Cell-to-cell spread of HSV-2 was more greatly affected by the lack of pUL21 than HSV-1, and strain-specific differences in the requirement for pUL21 in cell-to-cell spread were also noted. HSV-2 strain 186 lacking pUL21 was particularly crippled in both cell-to-cell spread and viral replication in non-complementing cells, in comparison to other HSV strains lacking pUL21, suggesting that the strict requirement for pUL21 by strain 186 may not be representative of the HSV-2 species as a whole. This work highlights CRISPR/Cas9 technology as a useful tool for rapidly constructing deletion mutants of alphaherpesviruses, regardless of background strain, and should find great utility whenever strain-specific differences need to be investigated. View Full-Text
Keywords: HSV; UL21; CRISPR/Cas9 HSV; UL21; CRISPR/Cas9

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Finnen, R.L.; Banfield, B.W. CRISPR/Cas9 Mutagenesis of UL21 in Multiple Strains of Herpes Simplex Virus Reveals Differential Requirements for pUL21 in Viral Replication. Viruses 2018, 10, 258.

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