pone.0236175.g007.tif (238.69 kB)

Inhibition of PKC or Rho protein kinase decreases Ad5 wild type replication in vitro.

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posted on 2020-07-22, 17:30 authored by Alice Liu, Cristhian J. Ildefonso, Wesley S. Bond, Mary Y. Hurwitz, Richard L. Hurwitz

Y79 cells were infected in a 24 well plate (2x105 cells/well) with Ad5WT (MOI: 250 genomes/cell) in the presence of DMSO (1:103 dilution) for 24 hours, or with DMSO (1:103 dilution), or with Gö6983 (1 μM), or with Y27632 (100 μM) for 48 hours. Cells were harvested by centrifugation at 500 g for 10 minutes and trypsinized, followed by a wash with PBS 1X. Cell pellets were then stored at -80°C until all samples were collected. Total DNA was extracted using the DNEasy Kit (QIAGEN, Inc) as per the manufacturer's protocol. Ad genomes and cellular genomes were quantified using 125 ng of each sample in an Ad Q-PCR assay as described in Materials and methods. Ad genome numbers are standardized to cellular genome copies in each sample. Values represent the average ± standard deviation. (n = 6 biological replicates, p<0.0001).