Expression of a foreign gene by recombinant canine distemper virus recovered from cloned DNAs.
A canine distemper virus (CDV) genomic cDNA clone and expression plasmids required to establish a CDV rescue system were generated from a laboratory-adapted strain of the Onderstepoort vaccine virus. In addition, a CDV minireplicon was prepared and used in transient expression studies performed to identify optimal virus rescue conditions. Results from the transient expression experiments indicated that minireplicon-encoded reporter gene activity was increased when transfected cell cultures were maintained at 32 rather than 37 degrees C, and when the cellular stress response was induced by heat shock. Applying these findings to rescue of recombinant CDV (rCDV) resulted in efficient recovery of virus after transfected HEp2 or A549 cells were co-cultured with Vero cell monolayers. Nucleotide sequence determination and analysis of restriction site polymorphisms confirmed that rescued virus was rCDV. A rCDV strain also was engineered that contained the luciferase gene inserted between the P and M genes; this virus directed high levels of luciferase expression in infected cells.
Parks
Christopher L
CL
Wyeth-Lederle Vaccines, Department of Viral Vaccine Research, 401 North Middletown Road, Pearl River, NY 10965, USA.
Wang
Hai-Ping
HP
Kovacs
Gerald R
GR
Vasilakis
Nikos
N
Kowalski
Jacek
J
Nowak
Rebecca M
RM
Lerch
Robert A
RA
Walpita
Pramila
P
Sidhu
Mohinderjit S
MS
Udem
Stephen A
SA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D006224:Cricetinae; D004279:DNA, Viral; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D004217:Distemper Virus, Canine / Q000235:genetics* / Q000302:isolation & purification; D004285:Dogs; D015870:Gene Expression; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins / Q000235:genetics; D011995:Recombination, Genetic; D012093:Replicon; D014407:Tumor Cells, Cultured; D014709:Vero Cells
D004279:DNA, Viral; D019590:Nucleocapsid Proteins; D008156:Luciferases; D012321:DNA-Directed RNA Polymerases