Figure 3
From: SF2/ASF binding region within JC virus NCCR limits early gene transcription in glial cells
Viral propagation of mutant JCV strains in PHFA cells. A. Schematic representation of JCV wild type and mutant genomes. B. Western blot analyzes of whole cell extracts from PHFA cells infected with JCV-Mad1-WT and mutants, JCV-Mad1-(1x98), and JCV-Mad1-ΔCR3-(1X73), using specific antibodies against LT-Ag, VP1, SF2/ASF and Agno protein. In lanes 7 and 8, whole cell extracts from uninfected cells were loaded as negative control. Western blot analyzes of same extracts with anti-Grb2 antibody was used as loading control. DPI depicts “day post-infection”. C. Southern blot analyses of replicated JCV genomic DNAs in parallel to protein samples in panel A. In lanes 1, 2 and 3, 2 ng of linearized Mad1-WT, Mad1-(1X98), and Mad1-ΔCR3-(1X73) were used as positive controls, respectively. In lane 4, DNA samples from uninfected cells were loaded as negative control. D. Q-PCR analyses of the JCV copy numbers in growth media of the infected PHFA cultures. Culture media was collected at 14 dpi, and was processed for the detection of viral particles by Q-PCR as described earlier [12, 16].