FIG. 5.

PFGE analysis of packaged genomic DNAs. (a) Monolayers of BHK cells were infected with wt HSV-1 or KUL25NS as indicated and incubated in either the presence (+) or absence (−) of 200 μg of PAA/ml. At 18 h p.i., the cells were fractionated into nuclei (N) and cytoplasm (C), and DNase-resistant DNA was prepared by a gentle method without phenol extraction. Samples were subjected to PFGE, and the gel was blotted and hybridized to ³²P-labeled pGX153 (which contains BamHI P). (b) Monolayers of BHK cells were infected with wt HSV-1 or KUL25NS and incubated for 17 h in either the presence (+) or absence (−) of 200 μg of PAA/ml. Duplicate samples of DNase-resistant nuclear DNA were resolved on a pulsed-field gel and blotted. The membrane was divided in two and hybridized to pBE1 or pST17, as indicated. The markers (M) are 5-kbp ladders (Bio-Rad), and the positions of the 5- and 70-kbp fragments are shown. The 152-kbp band corresponds to full-length HSV-1 DNA. (c) Monolayers of BHK cells were infected with KUL25NS, 25R22, wt HSV-1 strain 17 syn⁺ (wt 17), or wt HSV-1 strain KOS (wt KOS) as indicated and incubated for 17 h. The cells were harvested, embedded in agarose, lysed, and digested with proteinase K in situ. Samples were resolved on a pulsed-field gel, blotted, and hybridized to pGX2. The positions of the wells and of 152-kbp linear genomes are indicated.