FIG. 4.

Stability of recombinant Bac-AAV viruses. (A) Southern blot analysis of baculovirus genomes. One microgram of viral DNA prepared from the third viral passage was digested with EcoRV, fractionated on 1% agarose gel, transferred to a nylon membrane, and then hybridized with a probe specific for the transgene sequence. The expected bands of 4 and 10 kb are indicated by an arrow. (B) The same filter was stripped and hybridized with a probe specific for the gentamicin gene, which recognizes a 1.3-kb band. The intensity of the bands (indicated by arrows) in both panels A and B were quantified with a phosphorimager according to the manufacturer’s instructions. (C) Western blot detection of Rep isoforms expressed in Sf9 cells infected with different baculoviruses at an MOI of 10. At 2 days postinfection the total cell extracts were analyzed for the presence of Rep proteins. Rep78, Rep68, and Rep52 are indicated by an arrow.