vero-based ul37-complementing cell line bd45  (Johns Hopkins HealthCare)

Journal: Journal of Virology

Article Title: Herpes Simplex Virus 1 UL37 Protein Tyrosine Residues Conserved among All Alphaherpesviruses Are Required for Interactions with Glycoprotein K, Cytoplasmic Virion Envelopment, and Infectious Virus Production

doi: 10.1128/JVI.01202-16

Figure Lengend Snippet: Delineation of UL37 amino acids involved in infectious virus production. (A) Schematic of the UL37 protein depicting known functional domains and the approximate locations of nine single- and double-amino-acid replacements constructed to assess their role in infectious virus production. (B) Images of viral plaques produced in the transfection-infection complementation of the UL37-null virus. Vero cells were transfected with each plasmid. After 36 h, plasmid-transfected cells were infected with UL37-null virus at an MOI of 5 and virus stocks were collected at 24 hpi. Viral plaques were visualized by immunohistochemistry at 48 hpi (see Materials and Methods). The plasmids encoding mutations in the amino acids of UL37 that were able to complement the UL37-null virus are marked with a green plus, and the mutations which were not able to complement are marked by a red minus. (C) Viral titers obtained in the transfection-infection complementation assay. Titers of virus stocks collected at 24 hpi with the UL37-null virus were determined on the BD45 cell line, which expresses the UL37 protein in trans . A plasmid expressing the wild-type UL37 protein was used as a positive control. A second positive control consisted of mock transfection, followed by infection with wild-type HSV-1, while a UL20-expressing plasmid was used as a negative control.

Article Snippet: The Vero cell-based UL37-complementing cell line BD45 was a gift from Prashant Desai (Johns Hopkins University, Baltimore, MD).

Techniques: Virus, Functional Assay, Construct, Produced, Transfection, Infection, Plasmid Preparation, Immunohistochemistry, Expressing, Positive Control, Negative Control

Journal: Journal of Virology

Article Title: Herpes Simplex Virus 1 UL37 Protein Tyrosine Residues Conserved among All Alphaherpesviruses Are Required for Interactions with Glycoprotein K, Cytoplasmic Virion Envelopment, and Infectious Virus Production

doi: 10.1128/JVI.01202-16

Figure Lengend Snippet: Construction and characterization of UL37 mutant viruses. (A) Schematic representation of the UL37 mutant viruses constructed on the viral genomic background via double-Red mutagenesis on the HSV-1(F) and VC1 genomes cloned as a bacterial artificial chromosome. The approximate locations of the constructed mutations are shown. (B to E) Replication kinetics of wild-type and selected mutant viruses on Vero and BD45 cells. Viruses were adsorbed on nearly confluent monolayers of Vero or BD45 cells. Cells were infected with each virus at an MOI of 2. Virus stocks were collected at the indicated times, and titers were determined on both Vero and BD45 cells. Virus titers from three independent cultures were averaged, and the standard deviation was calculated for each time point.

Article Snippet: The Vero cell-based UL37-complementing cell line BD45 was a gift from Prashant Desai (Johns Hopkins University, Baltimore, MD).

Techniques: Mutagenesis, Construct, Clone Assay, Infection, Virus, Standard Deviation