Generation of recombinant lentogenic Newcastle disease virus from cDNA.
Recombinant lentogenic Newcastle disease virus (NDV) of the vaccine strain Clone-30 was reproducibly generated after simultaneous expression of antigenome-sense NDV RNA and NDV nucleoprotein, phosphoprotein and RNA-dependent RNA polymerase from plasmids transfected into cells stably expressing T7 RNA polymerase. For this purpose, the genome of Clone-30, comprising 15186 nt, was cloned and sequenced prior to assembly into a full-length cDNA clone under control of a T7 RNA polymerase promoter. Recombinant virus was amplified by inoculation of transfection supernatant into the allantoic cavity of embryonated specific-pathogen-free (SPF) chicken eggs. Two marker restriction sites comprising a total of five nucleotide changes artificially introduced into noncoding regions were present in the progeny virus. The recombinant NDV was indistinguishable from the parental wild-type virus with respect to its growth characteristics in cell culture and in embryonated eggs. Moreover, an intracerebral pathogenicity index of 0.29 was obtained for both viruses as determined by intracerebral inoculation of day-old SPF chickens, proving that the recombinant NDV is a faithful copy of the parental vaccine strain of NDV.
Römer-Oberdörfer
Angela
A
Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, D-17498 Insel Riems, Germany1.
Mundt
Egbert
E
Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, D-17498 Insel Riems, Germany1.
Mebatsion
Teshome
T
Intervet International B.V., Wim de Körverstraat 35, NL-5830 AA Boxmeer, The Netherlands2.
Buchholz
Ursula J
UJ
Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, D-17498 Insel Riems, Germany1.
Mettenleiter
Thomas C
TC
Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, D-17498 Insel Riems, Germany1.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002642:Chick Embryo; D019976:Cloning, Organism; D018076:DNA, Complementary / Q000737:chemistry; D008969:Molecular Sequence Data; D009522:Newcastle disease virus / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018076:DNA, Complementary