Evaluation of thymidine kinase and neomycin phosphotransferase II positive selection systems for recovery of genetically atypical and recombinant DNA vaccine viruses.
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.
Katz
J B
JB
Diagnostic Virology Laboratory, National Veterinary Services Laboratories, Ames, Iowa 50010.
Middle
L A
LA
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*
D004274:DNA, Recombinant; D005819:Genetic Markers; D014614:Vaccines, Synthetic; D014765:Viral Vaccines; D010770:Phosphotransferases; D013937:Thymidine Kinase; D019868:Kanamycin Kinase