Rescue of Newcastle disease virus from cloned cDNA using an RNA polymerase II promoter.
Arch Virol
; 156(6): 979-86, 2011 Jun.
Article
em En
| MEDLINE
| ID: mdl-21327786
ABSTRACT
A new system was developed to improve the efficiency and simplify the procedure of recovery of Newcastle disease virus (NDV) from cloned cDNA. A full-length cDNA clone of mesogenic NDV vaccine strain Mukteswar was assembled from five subgenomic cDNA fragments and cloned into a plasmid allowing transcription driven by cellular RNA polymerase II. The full-length viral cDNA was flanked by hammerhead ribozyme (HamRz) and hepatitis delta virus ribozyme (HdvRz) sequences, resulted in the synthesis of antigenomic RNA with exact termini. Without supplying T7 RNA polymerase, infectious NDV could be generated efficiently in some eukaryotic cell lines by simultaneous transcription of antigenomic RNA from the full-length plasmid and expression of NP, P and L proteins from helper plasmids introduced by cotransfection. The efficiency of recovery with the conventional T7 promoter system based on BRS-T7 cells and the cytomegalovirus (CMV) promoter system was compared, and the results demonstrate that the new system facilitates the generation of recombinant NDV and more efficient than the T7 rescue system using BRS-T7.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
RNA Polimerase II
/
Vírus da Doença de Newcastle
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Regiões Promotoras Genéticas
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Clonagem Molecular
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DNA Complementar
Limite:
Animals
Idioma:
En
Ano de publicação:
2011
Tipo de documento:
Article