Selective assay for herpes simplex viruses expressing thymidine kinase.
A technique for selecting herpes simplex viruses expressing the viral thymidine kinase (TK+) from a population of predominantly TK- viruses was developed. This was accomplished by infecting TK- cells and incubating the cultures under a liquid overlay medium containing methotrexate. Since the TK- cells survive in this medium for only a limited period of time, it was necessary to add fresh uninfected TK- cells 48 h after infection. The technique allowed the detection and quantitation of the TK+ virus fraction in mixtures of TK+ and TK- viruses where the TK+ fraction was present in frequencies as low as 10(-5). It was also used to estimate reversion frequencies and to obtain and analyze TK+ revertants from TK- mutant strains of herpes simplex virus type 1.
D002460:Cell Line; D003470:Culture Media; D006801:Humans; D008727:Methotrexate; D008828:Microbiological Techniques; D009154:Mutation; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D013937:Thymidine Kinase / Q000235:genetics*; D014779:Virus Replication
D003470:Culture Media; D013937:Thymidine Kinase; D008727:Methotrexate