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Journal of General Virology (1999), 80, 2987-2995.
© 1999 Society for General Microbiology

Animal: RNA Viruses

Generation of recombinant lentogenic Newcastle disease virus from cDNA

Angela Römer-Oberdörfer1, Egbert Mundt1, Teshome Mebatsion2, Ursula J. Buchholz1 and Thomas C. Mettenleiter1

Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, D-17498 Insel Riems, Germany1
Intervet International B.V., Wim de Körverstraat 35, NL-5830 AA Boxmeer, The Netherlands2

Author for correspondence: Angela Römer-Oberd örfer.Fax +49 38351 7219. e-mail angela.oberdoerfer{at}rie.bfav.de

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.




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