Acute replication of HSV-1 in SCG neurons follows the canonical ordered cascade of mRNA accumulation.

<p>(A) Primary neurons were isolated from superior cervical ganglia (SCG) of E21 rats, cultured for 7 days in the presence of 5 µM aphidicolin and 20 µM 5-fluorouracil to eliminate proliferating cells, and then infected with HSV-1 GFP-Us11 at a multiplicity of 3 plaque forming units per neuron (MOI = 3). RNA was collected at 0, 3, 6, 9, and 12 h post-infection (p.i.) and analyzed by quantitative reverse transcription PCR (qRT-PCR) to determine the relative levels of viral immediate-early (ICP27), early (UL30), γ1 leaky-late (VP16) and γ2 true-late (UL36) transcripts. Values represent the average and standard error from the mean from three independent infection experiments. (B) Neuron cultures were treated with the viral DNA polymerase inhibitor phosphonoacetic acid (PAA, 300 µg/ml) for 1 h (hatched bars) or mock treated (filled bars) and then infected with HSV-1 GFP-Us11. Total DNA was prepared at the indicated times and the relative levels of viral genomic DNA determined by quantitative (qPCR) using primers complementary to the HSV-1 UL30 gene. Input DNA was normalized by qPCR detection of the rat RPL19 gene. (C, D) Analysis of γ1 leaky-late (VP16) and γ2 true-late (UL36) transcript levels in the presence or absence of PAA.