RESCUE OF MEASLES VIRUSES FROM CLONED DNA

A system has been established allowing the rescue of replicating measl es viruses (MVs) from cloned DNA, On one hand, plasmids were construct ed from which MV antigenomic RNAs with the correct termini are transcr ibed by phage T7 RNA polymerase. On the other hand, helper cells deriv ed from the human embryonic kidney 293 cell line were generated consti tutively expressing T7 RNA polymerase together with MV nucleocapsid pr otein and phosphoprotein. Simultaneous transfection of the helper cell s with the MV antigenomic plasmid and with a plasmid encoding the MV p olymerase under direction of a T7 promoter led to formation of syncyti a from which MVs were easily recovered, A genetic tag comprising three nucleotide changes was present in the progeny virus, As a first appli cation of reverse genetics, a segment of 504 nucleotides from the 5' n on-coding region of the fusion gene was deleted,leading to an MV varia nt whose replication behaviour in Vero cells was indistinguishable fro m that of the laboratory Edmonston B strain, Since no helper virus is involved, this system, in principle, should be applicable to the rescu e of any member of the large virus order Mononegavirales, i.e. viruses with a non-segmented negative-strand RNA genome.