パルスフィールドグラディエント電気泳動法による単純ヘルペスウイルス感染細胞におけるウイルスDNAの解析

抄録

The DNA prepared from Vero cells infected with herpes simplex virus type 1 (HSV-1) was analyzed by pulsed field gradient gel electrophoresis (PFG). PFG was performed under three conditions using field inversion gel electrophoresis or orthogonal field alternation gel electrophoresis methods. Southern blot analysis with cloned HSV-1 DNA fragments was attempted for identification of HSV-DNA. The major part of HSV-DNA comigrated with HSV virion DNA, and was identified as HSV genomic DNA. Approximately 10% of HSV-DNA migrated a shorter distance than the HSV genomic DNA. The short migrating component consisted of one to three discrete bands depending on the condition of PFG. The molecular size of these bands were estimated to be more than 200 kilobase pairs. The short migrating component showed a tendency to comigrate with degradated cellular DNA, suggesting close association of the short migrating component with host cell DNA during viral replication. The mobility of the short migrating component was influenced by the treatment with several antiviral agents. Demonstration of the short migrating component of HSV-DNA by PFG may be direct evidence of the presence of high-molecular-weight replicative intermediates, and these observations are consistent with the “rolling circle model” in which the replicative intermediates are present as a concatemeric form.