Rescue of functional replication origins from embedded configurations in a plasmid carrying the adenovirus genome.
AUTOR(ES)
Hanahan, D
RESUMO
A variant of the adenovirus type 5 genome which lacks EcoRI sites has been cloned in a bacterial plasmid after the addition of EcoRI oligonucleotide linkers to its ends. Closed circular forms of the recombinant viral genome were not infectious upon their introduction into permissive eucaryotic cells. The linear genome released by digestion of the 39-kilobase recombinant plasmid (pXAd) with EcoRI produced infectious virus at about 5% of the level of wild-type controls. The viruses which arose were indistinguishable from the parental strain, and the normal termini of the viral genome had been restored. Marker rescue experiments demonstrate that provision of a DNA fragment with a normal viral end improves infectivity. When a small fragment carrying a wild-type left end (the 0 to 2.6% ClaI-B fragment) was ligated to ClaI-linearized pXAd, virus was produced with efficiencies comparable to a similar reconstitution of the two ClaI fragments of the wild-type genome. These viruses stably carry the left-end fragment at both ends, leaving the normal right end embedded in 950 base pairs of DNA. The embedded right origin is inactive. The consensus of the analyses reported here is that a free end is a necessary configuration for the sequences which make up the adenovirus origin of replication.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=368696Documentos Relacionados
- Rescue of the Adeno-Associated Virus Genome from a Plasmid Vector: Evidence for Rescue by Replication
- Involvement of topoisomerases in replication, transcription, and packaging of the linear adenovirus genome.
- Studies of adenovirus-SV40 hybrid viruses. IV. An adenovirus type 2 strain carrying the infectious SV40 genome.
- Hybridization of mRNA from adenovirus-transformed cells to segments of the adenovirus genome.
- Superinfection rescue of an integrated defective polyomavirus genome.