title: Evaluation of thymidine kinase and neomycin phosphotransferase II positive selection systems for recovery of genetically atypical and recombinant DNA vaccine viruses.
abstract: Dominant phenotypic marker cell culture selection systems were evaluated for the recovery of thymidine kinase positive (TK+), thymidine kinase negative (TK-), and neomycin phosphotransferase II positive (NPT II+) viruses. From 1 to 100 pfu of each marker-positive virus was diluted into 10(6) pfu of marker-negative background virus prior to selection. All three selection systems recovered 100 or fewer pfu of marked virus from the background population (10(-4) fractional level). In some instances, 1 to 10 pfu of marked virus were recovered (10(-5)-10(-6) fractional levels). TK+ and NPT II+ selections yielded nearly pure marker positive virus (98.8% and 99.5% respectively). Populations surviving TK- selection were significantly less pure (58.7%, P less than 0.05). Purity of recovered viruses was independent of the initial fractional level of marker-positive virus. The use of TK+, TK-, and NPT II+ selection systems is discussed in the context of purity testing and environmental surveillance of recombinant DNA viral vaccines.
authors.affiliation: Diagnostic Virology Laboratory, National Veterinary Services Laboratories, Ames, Iowa 50010.
authors.forename: J B
authors.initials: JB
authors.lastname: Katz
authors.forename: L A
authors.initials: LA
authors.lastname: Middle
mesh_terms: D004274:DNA, Recombinant; D005069:Evaluation Studies as Topic; D005819:Genetic Markers; D019868:Kanamycin Kinase; D010770:Phosphotransferases / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics; D014614:Vaccines, Synthetic / Q000302:isolation & purification*; D014616:Vaccinia virus / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D014765:Viral Vaccines / Q000302:isolation & purification*
chemical_list: D004274:DNA, Recombinant; D005819:Genetic Markers; D014614:Vaccines, Synthetic; D014765:Viral Vaccines; D010770:Phosphotransferases; D013937:Thymidine Kinase; D019868:Kanamycin Kinase