Method for induction of mutations in physically defined regions of the herpes simplex virus genome.
A procedure was developed for inducing mutations in isolated restriction enzyme fragments of herpes simplex virus type 1 (HSV-1) DNA with nitrous acid. The mutations were then transferred to the viral genome by genetic recombination during cotransfection of rabbit kidney cells with the mutagenized fragments and intact HSV-1 DNA. The HpaI restriction enzyme fragments LD, B, LG, I, and J were mutagenized. Temperature-sensitive mutants were found at frequencies of 1 to 5% among the progeny of the transfections. Syncytial mutants also were found at high frequency when fragment B or LD was used for mutagenesis. Fifteen of these mutants, 11 temperature sensitive and 4 syncytial, were used for further studies, including complementation analysis, DNA synthesis, and marker rescue. Marker rescue data presented here and in the accompanying publication (A. L. Goldin, R. M. Sandri-Goldin, M. Levine, and J. C. Glorioso, J. Virol. 38: 50-58, 1981) confirm the map position of some of the newly isolated mutants.
D004262:DNA Restriction Enzymes; D004279:DNA, Viral / Q000096:biosynthesis / Q000235:genetics; D005814:Genes, Viral; D005818:Genetic Engineering / Q000379:methods; D009154:Mutation; D009608:Nitrous Acid; D011995:Recombination, Genetic; D018139:Simplexvirus / Q000235:genetics*
D004279:DNA, Viral; D004262:DNA Restriction Enzymes; D009608:Nitrous Acid